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17 publications mentioning dme-mir-92b

Open access articles that are associated with the species Drosophila melanogaster and mention the gene name mir-92b. Click the [+] symbols to view sentences that include the gene name, or the word cloud on the right for a summary.

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[+] score: 428
Other miRNAs from this paper: dme-mir-92a, dme-mir-124
Some jigr1 Isoforms Are Direct Targets of miR-92a and miR-92b In VivomiRNAs regulate the expression of their target genes post-transcriptionally, mostly by binding to target sequences in the 3’UTR and mediating mRNA degradation and/or translation inhibition [24]. [score:15]
Upregulation of Jigr1 seems to play a role in nuclear Prospero expression in miR-92 [–/–] neuroblasts, since reducing Jigr1 expression eliminated this phenotype. [score:8]
Jigr1 Upregulation Leads to Premature Neuroblast Differentiation in miR-92 [–/–] FliesTo determine whether upregulation of jigr1contributes to the miR-92 mutant phenotype, we carried out genetic interaction experiments. [score:7]
Moreover, in order to assess whether jigr1 upregulation mimics the miR-92 [–/–] phenotype, we overexpressed jigr1 in larval brain neuroblasts (Fig 8C). [score:6]
However, deletion of site 3 eliminated the suppression, showing that jigr1 mRNA is a direct target of miR-92a and miR-92b (Fig 7B). [score:6]
In summary, our results reveal a local regulatory loop in which miR-92a and miR-92b are expressed in the jigr1 transcription unit and also work in concert to prevent premature differentiation of neuroblasts by limiting expression of the host gene. [score:6]
So, using the same ends-out gene targeting method [16], we targeted the miR-92b locus on the miR-92a [–/–] mutant background to generate flies lacking both miR-92a and miR-92b (named as miR-92 [–/–]). [score:5]
Expression of miR-92a or miR-92b in the miR-92 [–/–] background with Insc-GAL4 that drives gene expression in neuroblasts largely rescued the life span defect, confirming that this phenotype is indeed due to loss of miR-92a and miR-92b (S4 Fig). [score:5]
In miR-92a [-/-] L3 heads, miR-92b expression was increased by 30% and in miR-92b [-/-] L3 heads, miR-92a expression was increased by 40% (S1B Fig). [score:5]
Indeed, expression of miR-92a or miR-92b in miR-92 [–/–] mutants significantly reduced jigr1 expression in neuroblasts (Fig 6D and 6E). [score:5]
Prospero is also a predicted target of miR-92a and miR-92b (Targetscan). [score:5]
We were intrigued by the finding that miR-92 family and jigr1 show complementary expression pattern in the larval brain (Fig 2E), which raises the possibility that jigr1 mRNA might be a target of miR-92 family. [score:5]
Since all miR-92 [–/–] phenotypes in neuroblasts can be rescued by expression of miR-92a or miR-92b, we wanted to see if the elevated Jigr1 level could be suppressed too. [score:5]
In contrast, direct regulation of some jigr1 isoforms by miR-92a and miR-92b largely accounts for the observed complementary expression domains of jigr1 and these miRNAs (Figs 3 and 6), although we cannot completely rule out the possibility other mechanisms may also contribute to jigr1 repression. [score:5]
Overexpression of miR-92a or miR-92b suppressed luciferase activity from a reporter with intact binding sites but had no effect on a reporter containing mutated binding sites (Fig 7D). [score:5]
Drosophila miR-92a and miR-92b and Their Host Gene jigr1 Are Expressed in the Same Transcriptional Unit in Larval BrainmiR-92a and miR-92b have the same expression profile (Fig 1 and S2 Fig), which suggests that they may be co-transcribed. [score:5]
To determine whether miR-92a and miR-92b are co-expressed with jigr1, we analyzed jigr1 expression by northern blot in wild type and deletion lines. [score:5]
Genetic analysis in Drosophila showed that downregulation of jigr1 by intragenic miR-92a and miR-92b is required for neuroblast self-renewal, providing an example of the functional significance of miRNA–host gene interactions in animal development. [score:5]
In HEK 293T cells, co -expression of miR-92a and/or miR-92b with a luciferase reporter carrying the jigr1 long 3’UTR in HEK 293T cells significantly suppressed luciferase activity (Fig 7B). [score:5]
Prospero mRNA is not a direct target of miR-92a and miR-92b. [score:4]
Some jigr1 Isoforms Are Direct Targets of miR-92a and miR-92b In Vivo. [score:4]
Thus, prospero mRNA is not a direct target of miR-92a and miR-92b (S7B and S7C Fig). [score:4]
miR-92a and miR-92b Are Expressed in Neuroblasts of the Drosophila Larval BrainThe miR-92 family is evolutionarily conserved (S1A Fig) but its function in neural development in Drosophila is unknown. [score:4]
We couldn’t detect any significant downregulation of luciferase activity by either miR-92a or miR-92b. [score:4]
Indeed, limiting the Jigr1 level, either with the deficiency covering the jigr1 locus or by downregulating it in neuroblasts with UAS-jigr1 -RNAi, largely rescued reduced neuroblast numbers in miR-92 [–/–] flies (Fig 8A) and brain size (Fig 8B). [score:4]
To determine whether upregulation of jigr1contributes to the miR-92 mutant phenotype, we carried out genetic interaction experiments. [score:4]
During larval development, miR-92 family limits jigr1 expression in neuroblasts and is essential for maintenance of a neuroblast pool. [score:4]
Jigr1 upregulation is responsible for premature neuroblast differentiation in miR-92 [–/–] flies. [score:4]
1005264.g008 Fig 8 Jigr1 upregulation is responsible for premature neuroblast differentiation in miR-92 [–/–] flies. [score:4]
Jigr1 Upregulation Leads to Premature Neuroblast Differentiation in miR-92 [–/–] Flies. [score:4]
miR-92a expression level is mostly higher than that of miR-92b at different developmental stages (S1B Fig). [score:4]
Moreover, upregulation of jigr1 also contributed significantly to the life span defect in miR-92 [–/–] flies (S8 Fig). [score:4]
miR-92a and miR-92b regulate neuroblast self-renewal by inhibiting premature differentiation. [score:4]
Drosophila miR-92a and miR-92b and Their Host Gene jigr1 Are Expressed in the Same Transcriptional Unit in Larval Brain. [score:3]
miR-92a and miR-92b Maintain Neuroblast Self-Renewal by Inhibiting Premature Differentiation. [score:3]
If miR-92a and miR-92b were transcribed together, deleting the promoter region of miR-92a would prevent expression of both miRNAs. [score:3]
Taken together, our results suggest that miR-92 family maintains neuroblast self-renewal in part through limiting Jigr1 expression. [score:3]
However, in miR-92 [–/–] brains, Jigr1 is expressed at high levels, especially in neuroblasts (Fig 6D). [score:3]
Mutating the first two predicted binding sites did not abolish the suppression of luciferase activity by miR-92a and/or miR-92b. [score:3]
Although Del #38 and Del #4 are 12.5 kb and 6.6 kb upstream of miR-92b, respectively, the loss of miR-92b expression resulting from these deletions suggests that a single long transcript contains miR-92a and miR-92b. [score:3]
Both miR-92a and miR-92b were expressed in the optic lobe and central brain in wild type flies; this expression was absent in miR-92a [–/–] and miR-92b [–/–] flies (please see detailed generation and characterization of these mutant flies below), respectively (Fig 1B–1E, S2B and S2C Fig). [score:3]
Since miR-92a and miR-92b have an identical seed sequence and are similarly expressed, they might compensate for each other. [score:3]
These reductions were fully rescued by expression of either miR-92a or miR-92b with Insc-GAL4, suggesting miR-92a and miR-92b are required cell autonomously in neuroblasts (Fig 5A and S5B Fig). [score:3]
To determine which cell types express miR-92a and miR-92b, we analyzed third instar larval brains by RNA in situ hybridization. [score:3]
1005264.g007 Fig 7miR-92a and miR-92b target their host gene jigr1. [score:3]
HEK293T cells were transfected with wild type or mutant miR-92 sensor plasmids or empty psiCHECK-2, pSuper-GFP-pri-miR-92a, or pSuper-GFP-pri-miR-92b expression vectors (described above) or pSuper-GFp-miR-124 using Fugene6 (Promega). [score:3]
Enlarged view of dashed box shows expression of miR-92b in Dpn [+] neuroblasts. [score:3]
In 3–5% neuroblasts in miR-92 [–/–] brains, Prospero was localized in the nucleus and decreased the expression level of the neuroblast nuclear marker Dpn (S5E Fig). [score:3]
In miR-92 mutant clones, some Dpn [+] neuroblast cells also showed nuclear Prospero expression (Fig 5E and S6D Fig). [score:3]
Although miR-92a and miR-92b are transcribed together with the long jigr1 transcript, miR-92a and miR-92b are enriched in neuroblasts, which express Jigr1 at a low level. [score:3]
Consistent with this, miR-92b expression was also lost in Del #19 flies, which lack miR-92a and the coding region of jigr1 (Fig 3B). [score:3]
Since a high level expression of miR-92a and miR-92b is seen in neuroblasts, we examined larval brain neuroblasts in miR-92 [–/–] flies (S5A Fig). [score:3]
In HEK 293T cells, expression of this transcript containing the miR-92b stem loop (Fig 3D) increased the amount of mature miR-92b, confirming that miR-92b can be processed from the 3’UTR of long jigr1 transcript (Fig 3E). [score:3]
If an elevated level of Jigr1 causes premature differentiation of neuroblasts in miR-92 mutants, reducing Jigr1 levels in miR-92 mutants could suppress this phenotype. [score:3]
Our findings suggest that the progressive loss of neuroblasts in miR-92 [–/–] brains is due to premature differentiation of these cells resulted from cell-autonomous effect of loss of miR-92, as shown by ectopic expression of nuclear Prospero, decreased BrdU uptake, and reduced cell size. [score:3]
Here we found that miR-92a and miR-92b work in concert to restrict Jigr1 expression in Drosophila larval neuroblasts and thereby maintain the neuroblast pool. [score:3]
Bar graph shows normalized mean luciferase activity of cells transfected with miR-92 expression plasmid to that of cells transfected with empty plasmid from two independent experiments. [score:3]
Our results reveal a novel function of the miR-92 family and identify another negative feedback loop as an essential regulator in neural stem cell development. [score:3]
Generation of miR-92a [–/–], miR-92b [–/–], and miR-92 [–/–] FliesEnds-out gene targeting was performed as described. [score:3]
miR-92a and miR-92b target their host gene jigr1. [score:3]
Expression of miR-92a and miR-92b in third instar larval brains. [score:3]
Similarly, acute loss and gain of miR-92b function in mouse cortex showed that miR-92b restricts the generation of intermediate progenitor cells by suppressing Tbr2 [36]. [score:3]
Thus, jigr1 isoforms containing this 5’UTR can be regulated directly by miR-92a and miR-92b through its 5’UTR. [score:3]
Expression profile of miR-92a and miR-92b in third Instar larval brain. [score:3]
Moreover, overexpression of miR-92a or miR-92b by Pros-Gal4 in wild type background decreased jigr1 level in third instar larval brain (Fig 6A and S7A Fig). [score:3]
In the central brain, miR-92a and miR-92b preferentially co-expressed with the neuroblast markers Miranda (Mira) (Fig 1B–1E) and Deadpan (Dpn) (S2B Fig and Fig 2C). [score:3]
To generate miR-92a and miR-92b expression vectors, Drosophila pri-miR-92a and pri-miR-92b were amplified from genomic DNA and cloned into pSuper-GFP vector. [score:3]
Next, we analyzed miR-92 [–/–] brains for expression of Prospero, a homeodomain transcription factor that is localized in the cortex in self-renewing larval neuroblasts but transiently moves to the nucleus in quiescent or terminally differentiating neuroblasts [21, 22]. [score:3]
In neuroblasts, miR-92a and miR-92b were highly expressed as a single transcriptional unit also containing jigr1 coding region. [score:3]
miR-92a and miR-92b have the same expression profile (Fig 1 and S2 Fig), which suggests that they may be co-transcribed. [score:3]
To determine whether jigr1 is a functional target of miR-92 in vivo, we first analyzed jigr1 mRNA levels in control and miR-92 [–/–] third instar larval brains by. [score:3]
Moreover, miR-92b maintains asymmetric division of neural stem cells by restricting Tis21 expression in mouse neocortex [37]. [score:3]
To further confirm that jigr1 is a direct target of miR-92a and miR-92b, we used a reporter silencing assay. [score:3]
Indeed, in Del #38 and Del #4 flies, which lack the sequence upstream of miR-92a, expression of both miR-92a and miR-92b was lost (Fig 3B). [score:3]
1: w [1118]; 2: miR-92 [4/4]; 3: miR-92 [1/4]; 4: Def(3R)BSC321/miR-92 [4]; 5: Del #1. (C) Expression level of Jigr1 normalized to alpha-Tubulin. [score:3]
These results suggest that miR-92 directly regulates its host gene jigr1. [score:3]
Nucleotide sequence alignment and expression of miR-92a and miR-92b. [score:3]
3’ UTR -dependent, miR-92 -mediated restriction of Tis21 expression maintains asymmetric neural stem cell division to ensure proper neocortex size. [score:3]
Generation of miR-92a [–/–] and miR-92b [–/–] flies by ends-out gene targeting. [score:3]
1005264.g004 Fig 4Generation of miR-92a [–/–] and miR-92b [–/–] flies by ends-out gene targeting. [score:3]
As we showed previously (Fig 5D and S5C Fig), some miR-92 [-/-] mutant neuroblasts have nuclear Prospero expression. [score:3]
Bar graph shows normalized mean luciferase activity of cells transfected with miR-92 expression plasmid to that of cells transfected with control miRNA from three independent experiments. [score:3]
Jigr1 and miR-92 Have Complementary Expression Patterns. [score:3]
Deleting only the coding region of jigr1 in Del #7 also eliminates both miR-92a and miR-92b expression (Fig 3B). [score:3]
To determine whether neuroblast survival is defective in miR-92 [–/–] flies, we immunostained the cells for activated Caspase-3. No significant increase in caspase activation was detected (S5C Fig), and blocking apoptosis by expressing the anti-apoptotic protein P35 in neuroblasts did not rescue this phenotype (S5D Fig). [score:3]
miR-92a and miR-92b Are Expressed in Neuroblasts of the Drosophila Larval Brain. [score:3]
Bar graph shows normalized mean values of the luciferase activity of cells transfected with a miR-92 expression plasmid relative to that of cells transfected with control miRNA from three independent experiments. [score:3]
Through a genetic knockout of both miR-92a and miR-92b in Drosophila, which has not been done so far in any other mo del organism, we discovered a novel function for miR-92a and miR-92b in neuroblast self-renewal. [score:2]
In order to investigate whether prospero mRNA can be directly downregulated by miR-92a and miR-92b, we performed luciferase reporter assay. [score:2]
These results raise the possibility that miR-92 family may contribute to neuroblast development. [score:2]
Here we show that the gene encoding jing-interacting gene regulatory 1 (jigr1), a putative DNA -binding protein containing MADF domain with unknown function [13], hosts miR-92a in the intron and miR-92b in the 3’UTR. [score:2]
The miR-92 family is evolutionarily conserved (S1A Fig) but its function in neural development in Drosophila is unknown. [score:2]
Generation of miR-92a and miR-92b Single- and Double-Knockout Mutant FliesTo analyze the function of the miR-92 family in neurogenesis in Drosophila, we first generated miR-92a and miR-92b single mutants by ends-out homologous recombination in which 140 bp deletions were generated after mini white was removed by Cre/LoxP -mediated recombination (Fig 4A) [16]. [score:2]
Unlike the single -knockout flies, which have a normal life span, miR-92 [–/–] flies have a reduced life span; most die within a week (S4 Fig). [score:2]
We also uncover the functional significance of this intragenic miRNA–host gene interaction through genetic knockout of both miR-92a and miR-92b. [score:2]
Generation of miR-92a and miR-92b Single- and Double-Knockout Mutant Flies. [score:2]
MicroRNA-92b regulates the development of intermediate cortical progenitors in embryonic mouse brain. [score:2]
However, phenotypes caused by loss of both miR-92a and miR-92b in mammals have not been reported yet. [score:1]
We cloned the sequence containing the intact and the mutated binding sites to the 5’UTR of the luciferase gene and co -transfected it together with miR-92a and/or miR-92b and control miRNA. [score:1]
At 96 and 120 hr after larval hatching (ALH), there were about 100 neuroblasts in the central brain in wild type, miR-92a [–/–], and miR-92b [–/–] flies, as shown by staining with the neuroblast marker Dpn (Fig 5A, S5A and S5B Fig). [score:1]
Loss of miR-92a or miR-92b was confirmed by PCR (Fig 4B) and northern blot analysis (Fig 4C). [score:1]
Analysis of miR-92 [-/-] mutant neuroblasts in third instar larval brains. [score:1]
In order to generate Del #1, miR-92 [-/-] lines, which have one loxP site at miR-92a locus and two additional loxP sites at miR-92b locus, were used. [score:1]
S4 FigSurvival curve of male flies of the wild type (n = 100), miR-92a [–/–](n = 100), miR-92b [–/–](n = 100) and miR-92 [–/–](n = 100) mutants. [score:1]
The decreased number of neuroblasts in miR-92 [–/–] flies could reflect defects in neuroblast formation, cell survival or self-renewal. [score:1]
1: w [1118] (n = 10); 2: miR-92 [4/4] (n = 10); 3: miR-92 [1/4] (n = 8); 4: Insc-GAL4/UAS-miR-92a; miR-92 [4/4] (n = 7); 5: Insc-GAL4/UAS-miR-92a; miR-92 [1/4] (n = 9); 6: Insc-GAL4/UAS-miR-92b; miR-92 [1/4] (n = 10). [score:1]
The seed sequences of miR-92a and miR-92b are shown in blue. [score:1]
miR-92a and miR-92b are transcribed in the same transcriptional unit. [score:1]
Two short 3’UTRs contain one miR-92a/miR-92b binding site and the long 3’UTR contains three. [score:1]
For BrdU chase experiments, first clones were induced in hs-flp70, Tub-Gal4, UAS-mCD8: GFP; FRT82B, Tub-Gal80 / FRT82+ and, hs-flp70, Tub-Gal4, UAS-mCD8: GFP; FRT82B, Tub-Gal80/ FRT82B, miR-92 [4] larvae at 24 ± 6 hr ALH. [score:1]
For the Jigr1-5’UTR sensor, oligonucleotides containing wild type and mutated miR-92a and miR-92b binding sites were synthesized, annealed, and cloned at the 5’ of the Renilla luciferase gene using the NheI site in psiCHECK-2 vector. [score:1]
Generation of miR-92a [–/–], miR-92b [–/–], and miR-92 [–/–] Flies. [score:1]
Error bars show S. D. (C) Percentage of neuroblasts with diameter > 10 μm (black bar) and < 10 μm (grey bar) in wild type (n = 60) and miR-92 [–/–] (n = 60) mutant brains at 96 hr ALH. [score:1]
5’ and 3’ DIG-labeled LNA -modified miR-92a and miR-92b probes (Exiqon) were used as recommended by the manufacturer. [score:1]
Thus, it is unclear what causes the lethality phenotype in miR-92b mutant flies generated by Chen et al [17]. [score:1]
The seed sequences of miR-92a and miR-92b are the same and shown in blue and mutated nucleotides are shown in red. [score:1]
1: w [1118] (n = 12); 2: miR-92a [-/-]; 3: miR-92b [-/-]; 4: miR-92 [4/4] (n = 11); 5: miR-92 [1/4] (n = 16); 6: Insc-GAL4/UAS-miR-92a; miR-92 [4/4] (n = 10); 7: Insc-GAL4/UAS-miR-92b; miR-92 [4/4] (n = 12); 8: Insc-GAL4/UAS-miR-92a; miR-92 [1/4] (n = 8); 9: Insc-GAL4/UAS-miR-92b; miR-92 [1/4] (n = 8). [score:1]
RNA was probed for miR-92a, miR-92b, U6, Jigr1 (exon 6 probe), Jigr1 (long 3’UTR probe) and rp49. [score:1]
In Drosophila, the miR-92 family consists of miR-92a and miR-92b, which have the same seed sequence (miRbase, S1A Fig). [score:1]
To investigate this possibility, we generated a series of deletions at the jigr1 locus by FLP-FRT [15] or cre/loxP deletions (Fig 3A and S3A Fig) and analyzed expression of miR-92a and miR-92b by northern blot (Fig 3B). [score:1]
jigr1 transcripts were almost threefold more abundant in miR-92 [–/–] brains (p < 0.0001) (Fig 6A). [score:1]
Both miR-92a [–/–] and miR-92b [–/–] flies were viable and fertile and had no obvious phenotype, in contrast to a recent report showing miR-92b deletion mutant flies are larval lethal, in which both miR-92b and the jigr1 coding region are deleted [17]. [score:1]
In northern blot, miR-92a level seems to be higher in miR-92b [-/-] mutants and vice versa (Fig 4C). [score:1]
Elevated Jigr1 levels in miR-92 [-/-] brains. [score:1]
miR-92a: 5’-ATAGGCCGGGACAAGTGCAATG-3’; miR-92b: 5’-GCAGGCCGGGACTAGTGCAATT-3’; U6: 5’-CACGAATTTGCGTGTCATCCTT-3’. [score:1]
1: w [1118]; 2: miR-92 [4/4]; 3: miR-92 [1/4]. [score:1]
To analyze the function of the miR-92 family in neurogenesis in Drosophila, we first generated miR-92a and miR-92b single mutants by ends-out homologous recombination in which 140 bp deletions were generated after mini white was removed by Cre/LoxP -mediated recombination (Fig 4A) [16]. [score:1]
Almost 80% of wild type neuroblasts incorporated BrdU, versus only 50% of miR-92 [–/—]neuroblasts, suggesting that loss of miR-92a and miR-92b causes premature exit from the cell cycle (Fig 5D). [score:1]
RNA was probed for miR-92a (top), miR-92b (middle), and U6 (bottom). [score:1]
The UAS-dsRed-miR-92a vector was from the Drosophila RNAi Screening Center, and UAS-miR-92b was generated by amplifying the 500–base pair pri-miR-92b from genomic DNA and cloning it into the pTW vector. [score:1]
S6 FigAsymmetric cell division is not disrupted in miR-92 [-/-] neuroblasts. [score:1]
In order to determine if size asymmetric division is disrupted in miR-92 [-/-] mutants, we have stained the third instar larval brains with Miranda and phosphor-histone H3. [score:1]
Thus, control of neuroblast self-renewal by miR-92a and miR-92b is achieved by preventing their early differentiation. [score:1]
w [1118] (n = 10); miR-92 [4/4] (n = 10); miR-92 [1/4] (n = 8); Def(3R)BSC321/miR-92 [4] (n = 12); Insc-GAL4/UAS-Jigr1 [RNAi] ; miR-92 [1/4] (n = 8). [score:1]
S1 Fig(A) Sequence alignment of miR-92a and miR-92b mature sequences from different species (miRBase). [score:1]
Next we analyzed miR-92 phenotype using mosaic analysis of repressible cell marker (MARCM) method [23]. [score:1]
miR-92b, miR-92a, and control U6 probes were used. [score:1]
At 120 hr ALH, the reduction was ~23% in miR-92 [1/4] flies (p < 0.005) and ~30% in miR-92 [4/4] flies (p < 0.0001) (Fig 5A). [score:1]
In this study, we report an unusual genomic arrangement in which miR-92a and miR-92b are embedded in the intron and 3’UTR of the host gene jigr1, respectively. [score:1]
Neither the apical localization of aPKC nor the basal localization of Numb or Miranda was disrupted in miR-92 [–/–] neuroblasts (n = 30) (S6A and S6B Fig). [score:1]
S7 Fig(A) Staining of third instar larval brains of Pros-GAL4/+, Pros-GAL4/UAS-miR-92a and Pros-GAL4/UAS-miR-92b with Prospero (blue) and Jigr1 (red). [score:1]
UAS-miR-92a and UAS-miR-92b transgenic flies were generated by Rainbow Transgenics. [score:1]
However, miR-92 [–/–] flies had significantly fewer neuroblasts (Fig 5A and S5B Fig). [score:1]
4: Pros-Gal4/+, 5: Pros-Gal4/UAS-miR-92a; 6: Pros-Gal4/UAS-miR-92b. [score:1]
1005264.g001 Fig 1(A) Northern blot analysis of total RNA isolated from staged wild type flies (lanes 1–13) and miR-92 [–/–] mutant third instar larvae (lane 14). [score:1]
However, in miR-92 [–/–] mutants, neuroblasts were lost progressively, resulting in 18% decrease at 96 hr ALH and a further 30% decrease at 120 hr ALH (p < 0.0001) (Fig 5B). [score:1]
Based on the FlyBase, miR-92a and miR-92b are located on the right arm of chromosome 3; miR-92a resides in the first intron of jigr1, and miR-92b resides downstream of jigr1 coding region. [score:1]
In controls, almost all neuroblasts were >10 μm in diameter at 96 hr ALH; however, in miR-92 [–/–] mutants, 39% of neuroblasts had a reduced cell diameter (p < 0.0001) (Fig 5C). [score:1]
The short 3’UTR of jigr1 has no binding site for miR-92a or miR-92b, while the long 3’UTR has three predicted miR-92 binding sites (Fig 7A). [score:1]
S5 FigAnalysis of miR-92 [-/-] mutant neuroblasts in third instar larval brains. [score:1]
Thus, defective asymmetric cell division is unlikely to be the mechanism of neuroblast loss in miR-92 [–/–] larvae. [score:1]
At 24, 48 and 72 hr ALH, wild type and miR-92 [–/–] flies had equal numbers of neuroblasts, suggesting that the phenotype is not due to defect in neuroblast formation or failure of neuroblasts to re-enter the cell cycle (Fig 5B). [score:1]
MARCM clones were generated as described before by crossing hs-flp70, Tub-Gal4, UAS-mCD8: GFP; FRT82B, Tub-Gal80 flies with FRT82B flies to generate control clones and with FRT82B, miR-92 [4] flies to generate mutant clones. [score:1]
1: w [1118] (n = 17); 2: miR-92a [-/-] (n = 13); 3: miR-92b [-/-] (n = 12); 4: miR-92 [4/4] (n = 15); 5: miR-92 [1/4] (n = 20); 6: Insc-GAL4/UAS-miR-92a; miR-92 [4/4] (n = 8); 7: Insc-GAL4/UAS-miR-92b; miR-92 [4/4] (n = 8); 8: Insc-GAL4/UAS-miR-92a; miR-92 [1/4] (n = 16); 9: Insc-GAL4/UAS-miR-92b; miR-92 [1/4] (n = 15). [score:1]
However, unlike Drosophila, human and mouse miR-92a and miR-92b genes are not intragenic. [score:1]
w [1118] (n = 17); miR-92 [4/4] (n = 15); miR-92 [1/4] (n = 20); Def(3R)BSC321/miR-92 [4] (n = 17); Insc-GAL4/UAS-Jigr1 [RNAi] ; miR-92 [1/4] (n = 14). [score:1]
1005264.g006 Fig 6Elevated Jigr1 levels in miR-92 [-/-] brains. [score:1]
Indeed, although no obvious difference in body size was observed for adult flies, the size of the ventral nerve cord of miR-92 [–/–] adult flies is 15% smaller (p < 0.05), the brains were also smaller than those of control flies (Fig 5G and 5H), which is probably due to reduced proliferation (Fig 5D) and premature differentiation (Fig 5E) of neuroblasts that lead to reduced production of neurons and glial cells. [score:1]
We didn’t observe any size symmetric neuroblast division in mir-92 [-/-] mutant neuroblasts (n = 33) (S6C Fig). [score:1]
However, our flies homozygous for Del #1, in which both miR-92a/miR-92b and Jigr1 are deleted, are still adult viable. [score:1]
Asymmetric cell division is not disrupted in miR-92 [-/-] neuroblasts. [score:1]
Our results also suggest that miR-92b does not have its own promoter and is not intergenic but is located in the 3’UTR of a longer jigr1 transcript in the Drosophila larval brain. [score:1]
Next, we asked whether the long jigr1 transcript could give rise to miR-92b. [score:1]
Moreover, the size of miR-92 mutant clones was significantly reduced (Fig 5F). [score:1]
1: w [1118]; 2: miR-92 [4/4]; 3: miR-92 [1/4]; 4: Def(3R)BSC321/miR-92 [4]; 5: Del #1. (D) Single confocal sections of third instar larval brains of the indicated genotypes stained for Jigr1 (red) and Miranda (green). [score:1]
We first characterized the expression of miR-92a and miR-92b in Drosophila at different stages of development by northern blot analysis and miRNA Taqman assay. [score:1]
One is 203 bp long and the other is 1827 bp long and contains the miR-92b stem loop (S3B and S3C Fig). [score:1]
S8 FigSurvival curve of w [1118] (red), miR-92 [-/-] (green), Def(3R)BSC321/miR-92 [4] (orange), Def(3R)BSC321/miR-92 [1] (yellow), Elav-GAL4/UAS-jigr1 [RNAi] ;miR-92 [1/4] (blue), Del #1/miR-92 [1] (brown) flies. [score:1]
These results suggest that miR-92a and miR-92b share the same transcriptional unit with a jigr1 isoform that hasn’t been previously annotated. [score:1]
In BrdU chase experiment in MARCM clones, around 40% of the wild type neurons were BrdU [+] while only 23% of miR-92 [–/–] neurons were BrdU [+]. [score:1]
1005264.g003 Fig 3 miR-92a and miR-92b are transcribed in the same transcriptional unit. [score:1]
Consistent with the increase in transcript levels, Jigr1 protein was also substantially more abundant in miR-92 [–/–] larval brains (Fig 6B and 6C). [score:1]
The miR-92b probe recognizes endogenous miR-92b in HEK 293T cells. [score:1]
Jigr1 mRNA has three potential miR-92 binding sites in the alternatively spliced noncoding exon 2 (Figs 3A and 7C). [score:1]
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[+] score: 51
Other miRNAs from this paper: dme-mir-92a, dme-mir-34, dme-mir-9c, dme-let-7, dme-mir-312
To confirm that expression of the luciferase reporter in this system is not simply sensitive to the coexpresssion of any miRNA, the experiment was also performed using miR-92b instead of let-7. Using the parameters noted above, MovingTargets does not predict mir-92b to target ab. [score:9]
However, by relaxing these constraints to require a maximum free energy of miRNA/target hybridization of -12 kcal/mole at room temperature and a minimum of 6 out of 8 consecutive base pairs in the miRNA 5' end, MovingTargets predicts that mir-9c, mir-92a, mir-92b, and mir-312 (the latter three are closely related miRNAs) target ttk. [score:7]
When the 'number of target sites' parameter of MovingTargets was relaxed to two, rather than three, we found that miR-92b is predicted to target ab at two sites that satisfy the remaining strict biological constraints. [score:7]
Exogenous miR-92b can repress expression in the assay system of a reporter mRNA bearing the 3' UTR of a predicted miR-92b target (below), but has only a small effect on luc/ab expression, much less than the effect of let-7 (Fig. 1B). [score:6]
Although miR-92b did not produce the strong inhibitory effect of let-7, it did cause some inhibition. [score:5]
Expression from the luciferase/ab mRNA with no added miRNA is shown at left, set to a relative value of 1. The other bars indicate the results of coexpression with let-7 or miR-92b. [score:5]
In our experiments to validate the predicted miRNA targets we tested a miRNA, miR-92b, not predicted to interact with the ab 3' UTR. [score:3]
The CrebA mRNA is predicted to have three target sites for miR-92b and mir-312 (Table 1); these miRNAs are closely related, sharing the same nts in positions 2–9 and 15–21. [score:3]
One interpretation is that miR-92b weakly interacts with the ab 3' UTR, and that the weak interaction is sufficient for a weak regulatory effect. [score:2]
We tested mir-92b and mir-312 in the S2 cell assay, and both miRNAs repress expression of the luc/ttk reporter (Figure 2C). [score:2]
Each primary transcript was amplified by PCR from genomic DNA to generate fragments with the following sequence coordinates: let-7: 2L:18450072-18450291; mir-92b: 3R:21466427-21466673; mir-312: 2R:15647675-15647897; mir-34: 3R:5926642-5926792. [score:1]
For panels A and C, the vertical bars above the line indicate sites for miR-92b, while vertical bars below the line indicate sites for miR-312. [score:1]
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[+] score: 38
Other miRNAs from this paper: hsa-mir-33a, hsa-mir-92a-1, hsa-mir-92a-2, dme-mir-1, dme-mir-8, dme-mir-11, hsa-mir-34a, hsa-mir-210, dme-mir-184, dme-mir-275, dme-mir-92a, dme-mir-276a, dme-mir-277, dme-mir-33, dme-mir-281-1, dme-mir-281-2, dme-mir-34, dme-mir-276b, dme-mir-210, dme-bantam, dme-mir-309, dme-mir-317, hsa-mir-1-2, hsa-mir-184, hsa-mir-190a, hsa-mir-1-1, hsa-mir-34b, hsa-mir-34c, aga-bantam, aga-mir-1, aga-mir-184, aga-mir-210, aga-mir-275, aga-mir-276, aga-mir-277, aga-mir-281, aga-mir-317, aga-mir-8, aga-mir-92a, aga-mir-92b, hsa-mir-92b, hsa-mir-33b, hsa-mir-190b, dme-mir-190, dme-mir-957, dme-mir-970, dme-mir-980, dme-mir-981, dme-mir-927, dme-mir-989, dme-mir-252, dme-mir-1000, aga-mir-1174, aga-mir-1175, aga-mir-34, aga-mir-989, aga-mir-11, aga-mir-981, aga-mir-1889, aga-mir-1890, aga-mir-1891, aga-mir-190, aga-mir-927, aga-mir-970, aga-mir-957, aga-mir-1000, aga-mir-309, cqu-mir-1174, cqu-mir-281-1, cqu-mir-1, cqu-mir-275, cqu-mir-957, cqu-mir-277, cqu-mir-252-1, cqu-mir-970, cqu-mir-317-1, cqu-mir-981, cqu-mir-989, cqu-mir-1175, cqu-mir-276-1, cqu-mir-276-2, cqu-mir-276-3, cqu-mir-210, cqu-mir-92, cqu-mir-190-2, cqu-mir-190-1, cqu-mir-1000, cqu-mir-11, cqu-mir-8, cqu-bantam, cqu-mir-1891, cqu-mir-184, cqu-mir-1890, cqu-mir-980, cqu-mir-33, cqu-mir-2951, cqu-mir-2941-1, cqu-mir-2941-2, cqu-mir-2952, cqu-mir-1889, cqu-mir-309, cqu-mir-252-2, cqu-mir-281-2, cqu-mir-317-2, aga-mir-2944a-1, aga-mir-2944a-2, aga-mir-2944b, aga-mir-2945, aga-mir-33, aga-mir-980
In contrast, miR-92 expression was upregulated in WNV-infected Cx. [score:6]
The targets of miR-989 and miR-92 in mosquitoes are not yet known; however, several studies have examined expression of these miRNAs during development. [score:6]
quinquefasciatus while miR-92 is significantly upregulated. [score:4]
Given the dysregulation of miR-989 and miR-92 during WNV infection, it is interesting to speculate that the targets of these miRNAs may play roles in mediating flavivirus infection in the mosquito host. [score:4]
Culex miR-989 and miR-92 expression levels are altered during flavivirus infection. [score:3]
Notably, this pattern of miRNA expression for miR-989 and miR-92 is also found in deep sequencing reads of WNV-infected Cx. [score:3]
Differences in miR-989 and miR-92 expression levels are highlighted. [score:3]
Two miRNAs, miR-92 and miR-989, showed significant changes in expression levels following WNV infection. [score:3]
Five miRNAs, miR-1, miR-317, miR-277, miR-989, and miR-92 were sequenced >120 times and were readily detectable in total RNA isolated from Cx. [score:1]
We additionally investigated the effects of flavivirus infection on miRNA expression and found that miR-92 and miR-989 are significantly changed in response to WNV infection. [score:1]
Five miRNAs, miR-184, miR-275, miR-277, miR-276, and miR-92, were sequenced >500 times and were readily detectable in total RNA isolated from C7/10 cells (Figure 3A). [score:1]
Furthermore, in vertebrates, miR-92 is a member of the conserved miR-17-92 cluster and is associated with oncogenesis and increased cellular proliferation. [score:1]
aegypti mosquitoes; miR-989 is also present in the midgut while miR-92 is present in Ae. [score:1]
In the silkworm Bombyx mori, miR-92 is associated with embryogenesis, a stage of high cellular proliferation and differentiation [49]. [score:1]
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[+] score: 22
Mating causes a reduction in expression of mir-34, mir-92b and mir-988 and an increase in expression of mir-286 (Figure  3; Additional file 4). [score:5]
Females that overexpress mir-34 and mir-92b lay 34% and 37% fewer eggs after mating, whereas females that overexpress mir-286 and mir-988 lay 16% and 33% more eggs than controls (Figure  7). [score:5]
To test the connectivity of the computational networks (Figures  4, 5 and 6) and evaluate causality, we overexpressed six miRNAs for which overexpression lines were available (mir-281-1, mir-286, mir-34, mir-92b, mir-310 and mir-988) using the binary GAL4-UAS expression system [38] under a universal ubiquitin promoter. [score:5]
To assess causality between post-mating modulation of miRNA expression and oviposition, we measured egg laying after mating by control females and females overexpressing mir-286, mir-34, mir-92b, or mir-988. [score:3]
Number of eggs laid by five young mated females over 18 hours between control and mir-286, mir-34, mir-92b and mir-988 overexpression lines. [score:3]
We obtained UAS-mir-281-1, UAS-mir-286, UAS-mir-34, UAS-mir-92b, UAS-mir-988, mir-310 and the co-isogenic control lines from the Bloomington Stock Center. [score:1]
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5
[+] score: 17
Our analyses in the very early stages of embryo development (NFE and ED0) of B. germanica, T. castaneum, D. melanogaster and D. virilis indicated that miRNAs from MIR-276 and MIR-279 families are differentially expressed in short germ-band species, whereas those of MIR-92 family are differentially expressed in long germ-band species. [score:6]
MIR-92 miRNAs downregulate jigr1 in the larval brain of D. melanogaster, which is essential for maintaining neuroblast self-renewal [25]. [score:4]
Mir-92 is expressed in ED0 and ED1 of B. germanica, being one of the miRNAs highly adenylated at the 3′ end, which suggests that it is cleared in the MZT. [score:3]
PCA (Fig. 5b) showed this grouping to be mostly driven by the MIR-276, MIR-279 and MIR-92 families. [score:1]
PCA showed the main drivers of this grouping to again be MIR-279 (for B. germanica and T. castaneum) and MIR-92 (for D. virilis) (Fig. 6b). [score:1]
In the tobacco hawk moth Manduca sexta, Mir-92 is clearly overexpressed in embryos compared to larvae, pupae and adults [37]. [score:1]
These included the MIR-92 and MIR-10 families, which are also highly adenylated in early embryos of T. castaneum [17]. [score:1]
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[+] score: 6
However, our individual strains with miR-SP [GenII] constructs directed against miR-92 family members (miR-92a,b and miR-310/311/312/313) did not display significant lethality, despite the fact that other phenotypes can be detected with SPs directed against this family (see below). [score:3]
For example, a previous study using a different lethality assay and SP design reported viability defects following miR-92 competitive inhibition 28. [score:2]
The identical phenotype was observed for miR-310SP, another member of the miR-92 family (Fig. 2f), suggesting some degree of functional redundancy between endogenous members of this miRNA family. [score:1]
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[+] score: 6
Figure 6 Sequence and expression of Ame-mir-0005/mir-92b. [score:3]
Analysis of the genomic location and mir-0005 mature sequence indicates that this is likely to be a paralogue of mir-92b (Figure 6A). [score:1]
Alignment of mir-0008 and mir-0005/mir-92b pre-miRNAs. [score:1]
Alignment of sequence reads to Apis mir-92a, mir-92b-1 and mir-0005/mir-92b pre-miRNAs. [score:1]
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[+] score: 5
Phylogenetic analysis, target gene prediction and pathway analysis showed that, among the 13 conserved miRNAs (miR-1, miR-100, miR-10a, miR-124, miR-125, miR-184, miR-33, miR-34, miR-7, miR-9, miR-92a, miR-92b and miR-let7), several highly conserved miRNAs (miR-1, miR-7 and miR-34) targeted the same or similar genes leading to the same pathways in shrimp, fruit fly and human (Figure 3b). [score:5]
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[+] score: 5
The two control miRNAs, bantam-3p and miR-92b-3p, whose expression levels remained consistent across all samples according to RNA-seq data, also showed no significant changes as determined by qRT-PCR. [score:3]
Two miRNAs, bantam-3p and miR-92b-3p were chosen as controls as their levels remained the same in knockdown and parental samples. [score:2]
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[+] score: 5
This cluster is highly conserved in the Drosophila lineage (Marco et al. 2013b), although it may have originated in insects (Ninova et al. 2014), and is evolutionarily related with the (also maternal) mir-92a/mir-92b cluster (Lu et al. 2008; Ninova et al. 2014). [score:1]
The most abundant microRNAs in unfertilized eggs were produced by mir-92b, mir-184, the mir-310/mir-311/mir-312/mir-313 cluster, and bantam genes, which accounted for over half of the microRNA reads. [score:1]
Other sets of microRNAs that may contribute to transcript clearance during MZT are the mir-310 and mir-92 families. [score:1]
In concordance to the high-throughput sequencing analysis presented in Figure 1, bantam-3p, mir-311-3p, and mir-92-3p were more abundant in the unfertilized egg than in the developing embryo. [score:1]
Although the microRNA level varies substantially across biological replicates, the presence of seven of the maternal microRNAs here described is validated (bantam-3p, mir-311-3p, mir-92b-3p, mir-184-3p, mir-14-3p, mir-995-3p, and mir-9c-5p), although the levels of the latter two were relatively low. [score:1]
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[+] score: 5
Moreover, mir-92a in Drosophila is hosted inside an intron, whereas mir-92b is not, suggesting that the two microRNAs may not be part of the same transcript. [score:1]
Although the statistical support of our phylogenetic analysis is weak (low bootstrap values), the fact that there is only one copy in Daphnia pulex also supports an insect specific duplication of mir-92. [score:1]
However, mir-92a/mir-92b may be the product of independent duplications in different animal lineages, i. e., mir-92a/mir-92b of protostomes and deutoerostomes may not be orthologous clusters (Supplementary Figure S1). [score:1]
Collectively, 14 clusters (the majority of our data set) emerged within the insects (Figure 3), that is, the Melanogaster, Drosophila and insect lineages in Table 1. Two clusters are conserved among all metazoans: the mir-125/let-7/mir-100 and the mir-92a/mir-92b clusters. [score:1]
Only two clusters are conserved in all metazoans, mir-92a/mir-92b and mir-125/let-7/mir-100. [score:1]
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[+] score: 4
Other miRNAs from this paper: dme-mir-92a
We confirmed these findings using a double mutant for miR-92a and its paralogue miR-92b [47], which exhibits an even smaller naked valley (Fig 2). [score:1]
S1 Fig(A) Trichomes gained in the naked valley after loss of miR-92a and miR-92b have a similar morphology as trichomes on the more distal femur. [score:1]
1007375.g002 Fig 2Leg trichome patterns in miR-92a/ miR-92b mutants. [score:1]
Leg trichome patterns in miR-92a/ miR-92b mutants. [score:1]
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[+] score: 3
Among the highly expressed; bantam, miR-184, miR-81, miR-100, miR-92, miR-2766, miR-279 are listed in the top (S1 Table) and these findings are in concurrence with recent reports [19]. [score:3]
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[+] score: 3
Other miRNAs from this paper: hsa-let-7a-1, hsa-let-7a-2, hsa-let-7a-3, hsa-let-7b, hsa-let-7c, hsa-let-7d, hsa-let-7e, hsa-let-7f-1, hsa-let-7f-2, hsa-mir-17, hsa-mir-25, hsa-mir-92a-1, hsa-mir-92a-2, hsa-mir-105-1, hsa-mir-105-2, dme-mir-1, dme-mir-10, mmu-let-7g, mmu-let-7i, mmu-mir-1a-1, mmu-mir-124-3, mmu-mir-134, mmu-mir-10b, hsa-mir-10a, hsa-mir-10b, dme-mir-92a, dme-mir-124, mmu-let-7d, dme-let-7, hsa-let-7g, hsa-let-7i, hsa-mir-1-2, hsa-mir-124-1, hsa-mir-124-2, hsa-mir-124-3, hsa-mir-134, mmu-let-7a-1, mmu-let-7a-2, mmu-let-7b, mmu-let-7c-1, mmu-let-7c-2, mmu-let-7e, mmu-let-7f-1, mmu-let-7f-2, mmu-mir-92a-2, hsa-mir-1-1, mmu-mir-1a-2, mmu-mir-10a, mmu-mir-17, mmu-mir-25, mmu-mir-124-1, mmu-mir-124-2, mmu-mir-92a-1, hsa-mir-379, mmu-mir-379, mmu-mir-412, gga-let-7i, gga-let-7a-3, gga-let-7b, gga-let-7c, gga-mir-92-1, gga-mir-17, gga-mir-1a-2, gga-mir-124a, gga-mir-10b, gga-let-7g, gga-let-7d, gga-let-7f, gga-let-7a-1, gga-mir-1a-1, gga-mir-124b, gga-mir-1b, gga-let-7a-2, gga-let-7j, gga-let-7k, dre-mir-10a, dre-mir-10b-1, dre-mir-430b-1, hsa-mir-449a, mmu-mir-449a, dre-let-7a-1, dre-let-7a-2, dre-let-7a-3, dre-let-7a-4, dre-let-7a-5, dre-let-7a-6, dre-let-7b, dre-let-7c-1, dre-let-7c-2, dre-let-7d-1, dre-let-7d-2, dre-let-7e, dre-let-7f, dre-let-7g-1, dre-let-7g-2, dre-let-7h, dre-let-7i, dre-mir-1-2, dre-mir-1-1, dre-mir-10b-2, dre-mir-10c, dre-mir-10d, dre-mir-17a-1, dre-mir-17a-2, dre-mir-25, dre-mir-92a-1, dre-mir-92a-2, dre-mir-92b, dre-mir-124-1, dre-mir-124-2, dre-mir-124-3, dre-mir-124-4, dre-mir-124-5, dre-mir-124-6, dre-mir-430b-2, dre-mir-430b-3, dre-mir-430b-4, dre-mir-430b-6, dre-mir-430b-7, dre-mir-430b-8, dre-mir-430b-9, dre-mir-430b-10, dre-mir-430b-11, dre-mir-430b-12, dre-mir-430b-13, dre-mir-430b-14, dre-mir-430b-15, dre-mir-430b-16, dre-mir-430b-17, dre-mir-430b-18, dre-mir-430b-5, dre-mir-430b-19, dre-mir-430b-20, hsa-mir-412, hsa-mir-511, dre-let-7j, hsa-mir-92b, hsa-mir-449b, gga-mir-449a, hsa-mir-758, hsa-mir-767, hsa-mir-449c, hsa-mir-802, mmu-mir-758, mmu-mir-802, mmu-mir-449c, mmu-mir-105, mmu-mir-92b, mmu-mir-449b, mmu-mir-511, mmu-mir-1b, gga-mir-1c, gga-mir-449c, gga-mir-10a, gga-mir-449b, gga-mir-124a-2, mmu-mir-767, mmu-let-7j, mmu-let-7k, gga-mir-124c, gga-mir-92-2, gga-mir-449d, mmu-mir-124b, gga-mir-10c, gga-let-7l-1, gga-let-7l-2
One example is the mir-25, 92/mir-92b case, in which the mature sequences are almost identical while the rest of the sequences share little sequence similarity (See additional file 6). [score:1]
For human miRNAs with same id numbers, only 2 are separated in the consensus families, namely mir-92/mir-92b and mir-449/mir-449b, showing that most of the miRNA families are robust to the variation in the input of the PBC pipeline. [score:1]
We further examined the alignments for mir-92/92b and mir-449/449b. [score:1]
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[+] score: 2
Among the old microRNAs, we have the mir-92, mir-184 and mir-9 families, which are conserved even in chordates. [score:1]
As a matter of fact, ongoing work in the laboratory has shown that the microRNAs mir-92a and mir-92b, and the mir-310/mir-311/mir-312/mir-313 cluster are abundant in Drosophila unfertilized eggs [33]. [score:1]
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[+] score: 1
The SLS output contained five predictions with significant similarity to the HOM output (ame-mir-13a, ame-mir-276, ame-mir-305, ame-mir-92 and ame-mir-9a) and only two predictions with significant similarity to the top 25 MCE candidates, both of which were variants of C5152. [score:1]
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[+] score: 1
This analysis revealed that no endogenous MREs were removed, except for the miR-92 family following integration of the T7-MRE [mut] barcode, as intended (Supplementary Table 1). [score:1]
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