A microRNA array reveals extensive regulation of microRNAs during brain development

May combine two claims.

MiR-19b signal was strong in E12 and E13, but nearly undetectable in the adult.

“MiR-19b signal was strong in E12 and E13, but nearly undetectable in the adult.”

May combine two claims.

Two different precursors of ∼65 nt, pre-mir9/131-ch1 and pre-mir9/131-ch15, but not pre-mir9/131-ch5, are detected with probes specific for the loop regions.

“Two different precursors of ∼65 nt, pre-mir9/131-ch1 and pre-mir9/131-ch15, but not pre-mir9/131-ch5, are detected with probes specific for the loop regions (Fig. 3C).”

May combine two claims.

Some miRNAs were overrepresented in polysomes, whereas others were underrepresented.

“Some miRNAs were overrepresented in polysomes, whereas others are underrepresented.”

May combine two claims.

The difference in miR-9 and miR-131 expression between PS1(+/-) and PS1(-/-) brains was less evident at E13.5 and enhanced over development from E13.5 to E17.5.

“The difference in miR-9 and miR-131 expression between PS1(+/-) and PS1(-/-) brains was less evident at E13.5 and enhanced over development from E13.5 to E17.5.”

May combine two claims.

miR-9 and miR-131 are brain-enriched transcripts processed from both arms of the stem of a ∼65 nt common precursor.

“Two of these miRNAs, miR-9 and miR-131, are brain-enriched transcripts (Fig. 3A) processed from both arms of the stem of a ∼65 nt common precursor (Fig. 3B).”

May combine two claims.

Both miR-9/miR-131 precursors are more abundant at E14-E15 than at E21.

“Although miR-9 and miR-131 expression patterns demonstrate their maximal level at E21, both precursors are more abundant at E14-E15, suggesting developmental regulation of their maturation.”

May combine two claims.

After E21, the miR-131 signal decreased, and in adults it appeared just 3.4-fold higher than at E12.

“After E21, the miR-131 signal decreased, and therefore in adults it appeared just 3.4-fold higher than at E12.”

May combine two claims.

At all three loci the sequences of miR-9 and miR-131 are perfectly conserved, whereas their joining sequences are just moderately conserved at two loci and differ strongly at a chromosome 15 locus.

“At all three loci the sequences of miR-9 and miR-131 are perfectly conserved, whereas their joining sequences are just moderately conserved at two loci (chromosomes 1 and 5), and differ strongly at a chromosome 15 locus (Fig. 3B).”

May combine two claims.

The expression patterns of the nine miRNAs fell into categories including postnatal expression, prenatal expression, peak expression at E21, increasing during the embryonic period followed by stable postnatal expression, and gradually increasing expression.

“The expression patterns fell into four categories: (1) postnatal expression, for example, miR-128; (2) prenatal expression, for example, miR-19b; (3) peak expression at E21, for example, miR-9, miR-125b, miR-131, miR-178; (4) increasing during the embryonic period followed by stable expression in the postnatal period, for example, miR-124a and miR-266; (5) gradually increasing expression, for exam”

May combine two claims.

Overall, a stronger signal was observed in the mRNPs; however, the proportion of specific miRNAs in polysomes versus mRNPs differed widely.

“Overall, a stronger signal was observed in the mRNPs; however, the proportion of specific miRNAs in polysomes versus mRNPs differed widely.”

May combine two claims.

MiR-124a signal increased more than 13-fold from E12 to E21, after which it remained stable.

“MiR-124a signal increased more than 13-fold from E12 to E21, after which it remained stable.”

May combine two claims.

The average increase in the miR-131 signal from E12 to E13 was 4.2-fold, and 4.7 from E13 to E21.

“For example, the average increase in the miR-131 signal from E12 to E13 was 4.2-fold, and 4.7 from E13 to E21.”

Probably not a substantial claim: may not pass S3 (load-bearing).

tRNAs comigrate with a synthetic marker of 65 nt.

“tRNAs here comigrate with a synthetic marker of 65 nt.”

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

A sequence within DNA helicase SMBP2 transcript is nearly complementary to miR-103.

“A sequence within DNA helicase SMBP2 transcript is nearly complementary to miR-103.”

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

In addition to miR-124a, its precursor is detected in lanes 4 and 5, but not in lane 6.

“In addition to miR-124a, its precursor is detected in lanes 4 and 5, but not in a lane 6.”

May combine two claims.

MiR-9 and miR-131 are disregulated in PS1 null mice.

“MiR-9 and miR-131 are disregulated in PS1 null mice.”

• AssumptionRests on an assumption: PS1(+/-) heterozygous mice are phenotypically indistinguishable from wild-type animals.

Probably not a substantial claim: may not pass S3 (load-bearing).

miR-271 is expressed in brain as detected by Northern blot analysis.

“Although this molecule is expressed in brain as detected by Northern blot analysis, it may represent a different class of non-coding RNA and, therefore, was deleted from the list of miRNAs included on the array.”

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

The low-molecular-weight RNA fraction allowed detection of several highly expressed pre-miRNAs and two tRNAs spotted as controls.

“Nevertheless, we were able to detect several highly expressed pre-miRNA, as well as two tRNAs, which were spotted as controls.”

Probably not a substantial claim: may not pass S3 (load-bearing).

The greater reduction in miR-9 compared to miR-131 possibly indicates a difference in their stability or processing.

“possibly indicating a difference in their stability or processing.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Inactivation of C. elegans Dicer (dcr-1) causes heterochronic phenotypes similar to let-7 and lin-4 mutations.

“Inactivation of C. elegans Dicer (dcr-1) causes heterochronic phenotypes similar to let-7 and lin-4 mutations (Grishok et al. 2001).”

• cites Grishok et al. 2001 sourceFetching the cited paper…

Probably not a substantial claim: may not pass S3 (load-bearing).

Two C. elegans miRNAs, lin-4 and let-7, control developmental timing by forming imperfect base pairing with elements within the 3' UTR of target mRNAs and attenuating their translation.

“Two C. elegans miRNAs, lin-4 and let-7, control developmental timing by forming imperfect base pairing with elements within the 3Ј UTR of target mRNAs and attenuating their translation (Lee et al. 1993;Wightman et al. 1993;Olsen and Ambros 1999;Reinhart et al. 2000;Slack et al. 2000).”

• cites Lee et al. 1993 sourceNot checked — no full text of the source
• cites Wightman et al. 1993 sourceNot checked — no full text of the source
• cites Olsen and Ambros 1999 sourceNot checked — no full text of the source

Probably not a substantial claim: may not pass S3 (load-bearing).

In Arabidopsis, suppression of RNA silencing, which shares a common pathway with miRNA processing, results in developmental defects.

“In Arabidopsis, suppression of RNA silencing, which shares a common pathway with miRNA processing, results in developmental defects (Kasschau et al. 2003).”

• cites Kasschau et al. 2003 sourceFetching the cited paper…

Probably not a substantial claim: may not pass S3 (load-bearing).

Many miRNAs, often with highly conserved sequences, are present in the genomes of C. elegans, Drosophila, mice, and humans.

“Many miRNAs, often with highly conserved sequences, are present in the genomes of Caenorhabditis elegans, Drosophila, mice, and humans (Lagos-Quintana et al. 2001, 2002;Lau et al. 2001;Lee and Ambros 2001;Mourelatos et al. 2002;Dostie et al. 2003).”

• cites Lagos-Quintana et al. 2001, 2002Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

• cites Lau et al. 2001 sourceNot checked — no full text of the source
• cites Lee and Ambros 2001 sourceNot checked — no full text of the source

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

In plants, miRNAs have greater complementarity to their mRNA targets and are likely to trigger degradation of their targets similarly to small interfering RNAs.

“In plants, miRNAs have greater complementarity to their mRNA targets, and are likely to trigger the degradation of their targets similarly to the action of small interfering RNAs (siRNAs; Llave et al. 2002;Rhoades et al. 2002).”

• cites Llave et al. 2002 sourceNot checked — no full text of the source
• cites Rhoades et al. 2002Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

Probably not a substantial claim: may not pass S3 (load-bearing).

miR-9 is also down-regulated during development of the fly, the organism from which it was initially cloned.

“Interestingly, miR-9 is also down-regulated during development of the fly, the organism from which it was initially cloned (Lagos-Quintana et al. 2001).”

• cites Lagos-Quintana et al. 2001 sourceNot checked — no full text of the source

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

The failure of miR-9 and miR-131 to up-regulate in PS1(-/-) embryonic brain development may be due to impaired Notch signaling.

“The failure of miR-9 and miR-131 to up-regulate in PS1(-/-) embryonic brain development may be due to impaired Notch signaling.”

Classified as probably not a substantial claim.

Premature neuronal differentiation in the PS1(-/-) brain is associated with aberrant neuronal migration and disorganization of the laminar architecture of the developing cerebral hemisphere.

“Premature neuronal differentiation in the PS1(-/-) brain is associated with aberrant neuronal migration and disorganization of the laminar architecture of the developing cerebral hemisphere (Handler et al. 2000).”

• cites Handler et al. 2000 sourceNot checked — no full text of the source

May combine two claims.

These precursors' expression can be independently regulated during brain development, and it is possible that only one member of the pair is expressed from each locus.

“Therefore, these precursors expression can be independently regulated during brain development, and it is possible that only one member of the pair is expressed from each locus.”

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

miRNA expression is regulated as a function of time course in neuronal cultures, and several miRNAs have been observed in association with polyribosomes in primary neurons.

“miRNA expression is regulated as a function of time course in neuronal cultures and several miRNAs have been observed in association with polyribosomes in primary neurons (J.Kim, A.M. Krichevsky, Y. Grad, G.D. Hayes, K.S. Kosik, G.M. Church, and G. Ruvkun, in prep.).”

• cites J.Kim, A.M. Krichevsky, Y. Grad, G.D. Hayes, K.S. Kosik, G.M. Church, and G. Ruvkun, in prep.Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

Probably not a substantial claim: may not pass S3 (load-bearing).

In Drosophila, bantam miRNA stimulates cell proliferation and tissue growth.

“In Drosophila, bantam miRNA stimulates cell proliferation and tissue growth (Brennecke et al. 2003).”

• cites Brennecke et al. 2003 sourceFetching the cited paper…

May combine two claims.

The lack of PS1 leads to premature differentiation of neural progenitor cells, while neural proliferation and apoptotic cell death during neurogenesis are unaltered.

“Here, the lack of PS1 leads to premature differentiation of neural progenitor cells, while neural proliferation and apoptotic cell death during neurogenesis are unaltered (Handler et al. 2000).”

• cites Handler et al. 2000 sourceNot checked — no full text of the source

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

The Drosophila homolog of FMRP associates with Dicer and with miRNAs and colocalizes with translating ribosomes.

“Drosophila homolog of this protein associates with Dicer and with miRNAs and colocalizes with translating ribosomes (Caudy et al. 2002).”

• cites Caudy et al. 2002Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

Probably not a substantial claim: may not pass S3 (load-bearing).

Some miRNAs are organized in the genome as clusters sometimes separated by intervals as short as a few nucleotides.

“Some miRNAs are organized in the genome as clusters sometimes separated by intervals as short as a few nucleotides (Lagos-Quintana et al. 2001;Lau et al. 2001).”

• cites Lagos-Quintana et al. 2001 sourceNot checked — no full text of the source
• cites Lau et al. 2001 sourceNot checked — no full text of the source

Probably not a substantial claim: may not pass S3 (load-bearing).

The enzyme responsible for cleavage of miRNA hairpin precursors is Dicer.

“The enzyme responsible for this cleavage is Dicer (Hutvagner et al. 2001;Ketting et al. 2001).”

• cites Hutvagner et al. 2001 sourceNot checked — no full text of the source
• cites Ketting et al. 2001 sourceFetching the cited paper…

Classified as probably not a substantial claim. May combine two claims.

In C. elegans both lin-4 miRNA and its target lin-14 mRNA associate with polyribosomes and lin-4 inhibits the translation of lin-14 mRNA after initiation of its translation.

“In C. elegans both lin-4 miRNA and its target lin-14 mRNA associate with polyribosomes and lin-4 inhibits the translation of lin-14 mRNA after initiation of its translation (Ambros 2000).”

• cites Ambros 2000 sourceNot checked — no full text of the source

Classified as probably not a substantial claim. May combine two claims.

Heterochronic RNAs in C. elegans control cell cycle progression and a particular sequence of cell divisions that is necessary for the generation of a specific cell type.

“Heterochronic RNAs in C. elegans control cell cycle progression and a particular sequence of cell divisions that is necessary for the generation of a specific cell type (Ambros 2000).”

• cites Ambros 2000 sourceNot checked — no full text of the source

Probably not a substantial claim: may not pass S3 (load-bearing).

The SMN protein is part of a large ribonucleoprotein complex that contains numerous miRNAs.

“This protein is part of a large ribonucleoprotein complex that contains numerous miRNAs (Mourelatos et al. 2002).”

• cites Mourelatos et al. 2002 sourceFetching the cited paper…

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

Many miRNAs have been isolated from mammalian embryonic neurons and mature brain, and some of them are neuronal-specific.

“Many miRNAs have been isolated from mammalian embryonic neurons and mature brain, and some of them are neuronalspecific (Lagos-Quintana et al. 2002;Dostie et al. 2003).”

• cites Lagos-Quintana et al. 2002 sourceFetching the cited paper…
• cites Dostie et al. 2003 sourceFetching the cited paper…

Probably not a substantial claim: may not pass S3 (load-bearing).

Samples with abnormally low incorporation were excluded from the analysis.

“Samples with abnormally low incorporation were excluded from the analysis.”

Probably not a substantial claim: may not pass S3 (load-bearing).

The estimated number of total miRNAs in vertebrates is ∼250 molecules.

“The estimated number of the total miRNAs in vertebrates is ∼250 molecules (Lim et al. 2003).”

• cites Lim et al. 2003 sourceNot checked — no full text of the source

May combine two claims.

The mentioned mRNAs encode either transcription factors or, like calcineurin, may affect transcription indirectly.

“Similar to the majority of the predicted plant targets (Rhoades et al. 2002), the mentioned mRNAs encode either transcription factors or, like calcineurin, may affect transcription indirectly.”

• cites Rhoades et al. 2002Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

Probably not a substantial claim: may not pass S3 (load-bearing).

Expression ratios are shown only where the ratio exceeds twofold and the corresponding p-value is equal to or below 0.05.

“Expression ratios are shown only where the ratio exceeds twofold and the corresponding p-value is equal or below 0.05.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Calcineurin A, and particularly its ␤ isoform expression in developing brain, is regulated posttranscriptionally, most likely at the translational level.

“Calcineurin A, and particularly its ␤ isoform expression in developing brain is regulated posttranscriptionally, most likely at the translational level (Polli et al. 1991).”

• cites Polli et al. 1991 sourceNot checked — no full text of the source

Classified as probably not a substantial claim.

Hybridization signals that failed to exceed the average background value by more than three standard deviations were excluded from analysis.

“Hybridization signals that failed to exceed the average background value by more than three standard deviations were excluded from analysis.”

Classified as probably not a substantial claim. May combine two claims.

mRNP and polysomal RNAs were isolated from three independent fractionation experiments, and each LMW RNA sample was hybridized in triplicate to the array membranes.

“mRNP and polysomal RNAs were isolated from three independent fractionation experiments, and each LMW RNA sample was hybridized in triplicate to the array membranes as shown in A.”

Classified as probably not a substantial claim.

The bars represent an average of three completely independent experiments.

“The bars represent an average of three completely independent experiments.”

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

The standard deviation between triplicate samples was <0.6 for all probes, <0.3 for 80% of probes, and <0.2 for 50% of probes.

“The standard deviation between triplicate samples was distributed as follows: <0.6 for all probes, <0.3 for 80% of probes, and <0.2 for 50% of probes.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Typically, an experiment included three independent RNA samples for each developmental time point.

“Typically, an experiment included three independent RNA samples for each developmental time point.”

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

Otx1 may represent another putative target for miR-103.

“and may represent another putative target for miR-103.”

Probably not a substantial claim: may not pass S3 (load-bearing).

miR-131 is alternatively known in the Rfam database as miR-9*, as it appears to be expressed from the opposite strand of the miR-9 precursor.

“In addition, miR-131 is alternatively known in the Rfam database as miR-9*, as it appears to be expressed from the opposite strand of the miR-9 precursor.”

• Also stated in: § Additional information · plain

  “Similarly, miR-131 is alternatively named miR-9*, as it is expressed from the opposite strand of the miR-9 precursor.”

• cites RfamNot matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

Classified as probably not a substantial claim. May combine two claims.

After normalization, data were averaged among duplicate arrays, and the resulting three data sets, each corresponding to an RNA sample, were treated as independent measurements for a two-tailed, two-sample t-test comparing developmental stages.

“Upon normalization, the remaining data was averaged among duplicate arrays, and the resulting three data sets each corresponding to an RNA sample were considered independent measurements for the purposes of the two-tailed, two-sample t-test when comparing different developmental stages.”

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

Mutation of SMBP2 causes mouse neuromuscular degeneration, a disease similar to human spinal muscular atrophy and linked to miRNA-containing complex.

“Mutation of this protein causes mouse neuromuscular degeneration (Cox et al. 1998), a disease similar to human spinal muscular atrophy and linked to miRNA-containing complex (see above).”

• cites Cox et al. 1998 sourceFetching the cited paper…

Probably not a substantial claim: may not pass S3 (load-bearing).

miRNAs miR-322-miR-351 have been recently cloned from rat neurons by Kim et al. (2003).

“miRNAs miR-322-miR-351 have been recently cloned from rat neurons by Kim et al. (2003).”

• cites Kim et al. (2003)Not matched

  Couldn’t be matched to a paper in Crossref, so it can’t be checked.

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

Many tiny noncoding RNA molecules that are not processed from a miRNA-like hairpin precursor and are not phylogenetically conserved have been recently cloned from C. elegans and are likely to be expressed in vertebrates as well.

“In addition, many tiny noncoding RNA molecules that are not processed from a miRNA-like hairpin precursor and are not phylogenetically conserved have been recently cloned from C. elegans and likely to express in vertebrates as well (Ambros et al. 2003).”

• cites Ambros et al. 2003 sourceFetching the cited paper…

Probably not a substantial claim: may not pass S3 (load-bearing).

To ensure accuracy of the hybridizations, each RNA sample was hybridized with two or three membranes.

“To ensure accuracy of the hybridizations, each RNA sample was hybridized with two or three membranes.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Otx1 is necessary for normal corticogenesis.

“Otx1, murine homolog of the Drosophila transcription factor orthodenticle, is necessary for normal corticogenesis (Simeone et al. 2002), and may represent another putative target for miR-103.”

• cites Simeone et al. 2002 sourceNot checked — no full text of the source

Probably not a substantial claim: may not pass S3 (load-bearing).

5S rRNA was detected by ethidium bromide staining to verify equal loading of total RNA.

“5S rRNA was detected by ethidium bromide staining of the gels prior to transfer to verify equal loading of total RNA.”