Most Caenorhabditis elegans microRNAs Are Individually Not Essential for Development or Viability

“We performed a broad phenotypic study of all available miRNA loss-of-function mutants, including mutants that had been reported earlier [2,4,20,52].”

“]. Because many messages have been under selective pressure to preserve pairing to a 6mer in the 5′ region of the miRNA known as the miRNA seed (nucleotides 2–7), targets of metazoan miRNAs can be predicted above the background of false-positives by searching for conserved matches to the seed region [”

“In animals, miRNAs are transcribed as long RNA precursors (pri-miRNAs), which are processed in the nucleus by the RNase III enzyme complex Drosha-Pasha/DGCR8 to form the approximately 70-base pre-miRNAs [21–25] or are derived directly from introns [26,27].”

“Because many messages have been under selective pressure to preserve pairing to a 6mer in the 59 region of the miRNA known as the miRNA seed (nucleotides 2–7), targets of metazoan miRNAs can be predicted above the background of false-positives by searching for conserved matches to the seed region [41–45].”

“miRNAs have been identified in organisms as diverse as roundworms, flies, fish, frogs, mammals, flowering plants, mosses, and even viruses, using genetics, molecular cloning, and predictions from bioinformatics [7–16].”

“We performed a broad phenotypic study of all available miRNA loss-of-function mutants, including mutants that had been reported earlier [2,4,20,52].”

“Because many messages have been under selective pressure to preserve pairing to a 6mer in the 59 region of the miRNA known as the miRNA seed (nucleotides 2–7), targets of metazoan miRNAs can be predicted above the background of false-positives by searching for conserved matches to the seed region [41–45].”

“elegans miRNAs [87,88].”

“Knockdown of miRNAs using modified 29 O-methyl antisense oligoribonucleotides also has been reported for the mouse [80].”

“elegans affects vulval development [89,90].”

“elegans miRNAs were cloned and verified in northern blot experiments [10,11,17,85].”

“For example, the miRNA miR-1 is expressed in muscles of Drosophila, the zebrafish, and the mouse [11,56,98].”

“In animals, miRNAs are transcribed as long RNA precursors (pri-miRNAs), which are processed in the nucleus by the RNase III enzyme complex Drosha-Pasha/DGCR8 to form the approximately 70-base pre-miRNAs [21–25] or are derived directly from introns [26,27].”

“elegans and mouse development [67,76].”

“In animals, miRNAs are transcribed as long RNA precursors (pri-miRNAs), which are processed in the nucleus by the RNase III enzyme complex Drosha-Pasha/DGCR8 to form the approximately 70-base pre-miRNAs [21–25] or are derived directly from introns [26,27].”

“Pre-miRNAs are exported from the nucleus by Exportin-5 [28], processed by the RNase III enzyme Dicer, and incorporated into an Argonaute-containing RNA-induced silencing complex (RISC) [29].”

“elegans and humans [94], appears not to have been an exception: for 15 miRNA families, miRNAs with identical seeds have been found in C.”

“Because many messages have been under selective pressure to preserve pairing to a 6mer in the 59 region of the miRNA known as the miRNA seed (nucleotides 2–7), targets of metazoan miRNAs can be predicted above the background of false-positives by searching for conserved matches to the seed region [41–45].”

“In animals, miRNAs are transcribed as long RNA precursors (pri-miRNAs), which are processed in the nucleus by the RNase III enzyme complex Drosha-Pasha/DGCR8 to form the approximately 70-base pre-miRNAs [21–25] or are derived directly from introns [26,27].”

“The first miRNA studied functionally in Drosophila is encoded by the bantam locus, which had previously been identified in a screen for deregulated tissue growth [54].”

“elegans [66,67], Drosophila [68,69], the zebrafish [70,71], and the mouse [72–75].”

“The first loss-of-function studies of miRNAs in the mouse have been reported demonstrating a role for miR-1 and miR208 in cardiac growth in response to stress [61,62] and miR155/BIC in normal immune function [63,64].”

“miRNAs have been identified in organisms as diverse as roundworms, flies, fish, frogs, mammals, flowering plants, mosses, and even viruses, using genetics, molecular cloning, and predictions from bioinformatics [7–16].”

“Binding of the silencing complex causes translational repression [31–33] and/or mRNA destabilization, which is sometimes through direct mRNA cleavage [34,35], but usually through other mechanisms [36–40].”

“elegans [66,67], Drosophila [68,69], the zebrafish [70,71], and the mouse [72–75].”

“For example, the miRNA miR-1 is expressed in muscles of Drosophila, the zebrafish, and the mouse [11,56,98].”

“The first loss-of-function studies of miRNAs in the mouse have been reported demonstrating a role for miR-1 and miR208 in cardiac growth in response to stress [61,62] and miR155/BIC in normal immune function [63,64].”

“elegans [66,67], Drosophila [68,69], the zebrafish [70,71], and the mouse [72–75].”

“For example, the severity of the muscle defect of C. elegans mir-1 mutants might depend on physiological conditions, as is the case for the Dmir-1 mutant phenotypes of Drosophila [56].”

“elegans miRNAs were cloned and verified in northern blot experiments [10,11,17,85].”

“]. Binding of the silencing complex causes translational repression [”

“miRNAs have been identified in organisms as diverse as roundworms, flies, fish, frogs, mammals, flowering plants, mosses, and even viruses, using genetics, molecular cloning, and predictions from bioinformatics [7–16].”

“Binding of the silencing complex causes translational repression [31–33] and/or mRNA destabilization, which is sometimes through direct mRNA cleavage [34,35], but usually through other mechanisms [36–40].”

“In animals, miRNAs are transcribed as long RNA precursors (pri-miRNAs), which are processed in the nucleus by the RNase III enzyme complex Drosha-Pasha/DGCR8 to form the approximately 70-base pre-miRNAs [21–25] or are derived directly from introns [26,27].”

“A second Drosophila miRNA, miR-14, also reduces programmed cell death [55].”

“elegans genome [93], about 60% of miRNAs are members of a family of two to eight genes [19].”

“miRNAs have been identified in organisms as diverse as roundworms, flies, fish, frogs, mammals, flowering plants, mosses, and even viruses, using genetics, molecular cloning, and predictions from bioinformatics [7–16].”

“In animals, miRNAs are transcribed as long RNA precursors (pri-miRNAs), which are processed in the nucleus by the RNase III enzyme complex Drosha-Pasha/DGCR8 to form the approximately 70-base pre-miRNAs [21–25] or are derived directly from introns [26,27].”

“Binding of the silencing complex causes translational repression [31–33] and/or mRNA destabilization, which is sometimes through direct mRNA cleavage [34,35], but usually through other mechanisms [36–40].”

“elegans [66,67], Drosophila [68,69], the zebrafish [70,71], and the mouse [72–75].”

“Pre-miRNAs are exported from the nucleus by Exportin-5 [28], processed by the RNase III enzyme Dicer, and incorporated into an Argonaute-containing RNA-induced silencing complex (RISC) [29].”

“Binding of the silencing complex causes translational repression [31–33] and/or mRNA destabilization, which is sometimes through direct mRNA cleavage [34,35], but usually through other mechanisms [36–40].”

“miRNA function has also been inferred from studies in which miRNAs have been misexpressed in worms, flies, frogs, mice, and cultured mammalian cells [65].”

“during studies of the control of developmental timing [”

“We performed a broad phenotypic study of all available miRNA loss-of-function mutants, including mutants that had been reported earlier [2,4,20,52].”

“The let-7 miRNA controls hypodermal cell-fate decisions during late-larval development by regulating the lin-41, hbl-1, daf-12, and pha-4 mRNAs [48–51].”

“elegans miRNAs were cloned and verified in northern blot experiments [10,11,17,85].”

“Within the silencing complex, metazoan miRNAs pair to the mRNAs of protein-coding genes, usually through imperfect base-pairing with the 39-UTR, thereby specifying the posttranscriptional repression of these target mRNAs [6,30].”

“For example, the severity of the muscle defect of C. elegans mir-1 mutants might depend on physiological conditions, as is the case for the Dmir-1 mutant phenotypes of Drosophila [56].”

“elegans [66,67], Drosophila [68,69], the zebrafish [70,71], and the mouse [72–75].”

“elegans [78,79] and appears to offer sequence-specific inhibition of small RNAs for a limited time span.”

“miRNAs have been identified in organisms as diverse as roundworms, flies, fish, frogs, mammals, flowering plants, mosses, and even viruses, using genetics, molecular cloning, and predictions from bioinformatics [7–16].”

“elegans [78,79] and appears to offer sequence-specific inhibition of small RNAs for a limited time span.”

“elegans [66,67], Drosophila [68,69], the zebrafish [70,71], and the mouse [72–75].”

“elegans [66,67], Drosophila [68,69], the zebrafish [70,71], and the mouse [72–75].”

“We previously described our studies of knockouts of three additional miRNA genes [52], and deletions in two other miRNA genes had been obtained by the C.”

“However, the predicted mRNA targets of miRNAs might not share the same degree of conservation as miRNA expression patterns—the spectrum of predicted mRNA targets varies significantly among metazoans [99].”

“elegans [66,67], Drosophila [68,69], the zebrafish [70,71], and the mouse [72–75].”

“The muscle-specific Drosophila miRNA miR-1 is required for larval development and cardiac differentiation [56,57].”

“Binding of the silencing complex causes translational repression [31–33] and/or mRNA destabilization, which is sometimes through direct mRNA cleavage [34,35], but usually through other mechanisms [36–40].”

“In nematodes, at least 10% of the protein-coding messages appear to be conserved targets of miRNAs [46].”

“Binding of the silencing complex causes translational repression [31–33] and/or mRNA destabilization, which is sometimes through direct mRNA cleavage [34,35], but usually through other mechanisms [36–40].”

“If let-60 RAS is a target of miR-84, loss of mir-84 might result in let-60 RAS overexpression and possibly a multivulva phenotype [91,92].”

“The let-7 miRNA controls hypodermal cell-fate decisions during late-larval development by regulating the lin-41, hbl-1, daf-12, and pha-4 mRNAs [48–51].”

“The let-7 miRNA controls hypodermal cell-fate decisions during late-larval development by regulating the lin-41, hbl-1, daf-12, and pha-4 mRNAs [48–51].”

“Injection of individual 29 O-methyl antisense oligoribonucleotides complementary to the 46 miRNAs known to be expressed in the fly embryo resulted in a total of 25 different abnormal phenotypes, including defects in patterning, morphogenesis, and cell survival [77].”

“elegans, flies, fish, and mammals, and several additional families are predicted to be conserved throughout these diverse lineages [19,95–97].”

“For example, genetic data indicate that Drosophila mir-7 directly regulates the transcriptional repressor Yan in the fly eye, but that loss ofmir-7 does not appreciably alter eye development, probably because of redundant protein turnover mechanisms that can also downregulate Yan [58].”

“Finally, Drosophila miR-9a is required for sensory organ precursor specification [59], and Drosophila miR-278 is required for energy homeostasis [60].”

“(2) Pharyngeal pumping: Frequency of grinder displacement was counted for 20 s by eye, but otherwise as described previously [102] (n 1⁄4 5).”

“Deletion mutants were isolated from a frozen library of worms mutagenized with ethyl methanesulphonate (EMS), 1,2,3,4-diepoxybutane (DEB), or a combination of UV irradiation and thymidine monophosphate (UV-TMP) [82,83].”

“Some miRNAs were predicted based on pre-miRNA folds and verified using northern blotting or PCR with specific primers and cloned miRNA libraries [17,18,85,86].”

“miRNAs have been identified in organisms as diverse as roundworms, flies, fish, frogs, mammals, flowering plants, mosses, and even viruses, using genetics, molecular cloning, and predictions from bioinformatics [7–16].”

“Finally, Drosophila miR-9a is required for sensory organ precursor specification [59], and Drosophila miR-278 is required for energy homeostasis [60].”

“Deletion mutants were isolated from a frozen library of worms mutagenized with ethyl methanesulphonate (EMS), 1,2,3,4-diepoxybutane (DEB), or a combination of UV irradiation and thymidine monophosphate (UV-TMP) [82,83].”

“elegans, flies, fish, and mammals, and several additional families are predicted to be conserved throughout these diverse lineages [19,95–97].”

“the control of developmental timing and likely regulate the hbl-1 mRNA, but act earlier in development than the let-7 miRNA [52,53].”

“org/ [107].”

“We made extensive use of the ‘‘poison’’ primer method, which increases the sensitivity of detection of small deletions [84].”

“Of the 95 C. elegans miRNA genes tested, only a few have grossly abnormal phenotypes when individually knocked out (Miska et al., 2007).”

“…Importance of miRNAs Systematic generation and analysis of miRNA knockouts in C. elegans created the initial impression that most metazoan miRNAs—even most miRNAs that are broadly conserved— have only subtle or redundant biological functions (Miska et al., 2007; Alvarez-Saavedra and Horvitz, 2010).”

“elegans coupled with in-depth screens for altered morphology, growth, development, and behavior identified only four gene knockouts with detectable phenotypes (Miska et al., 2007).”

“and Human Disease”

“In worms, most individual miRNA mutants show no gross phenotype (Miska et al., 2007); the same is true for several of the mouse knockouts generated to date, including miR-21, miR-210, miR-214, miR-206, and miR-143 (E. Olson, personal communication).”