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Genome-wide analysis of leafbladeless1-regulated and phased small RNAs underscores the importance of the TAS3 ta-siRNA pathway to maize development.

Dotto MC, Petsch KA, Aukerman MJ, Beatty M, Hammell M, Timmermans MC - PLoS Genet. (2014)

Bottom Line: Interestingly, in contrast to data from other plant species, we found no evidence for the existence of phased siRNAs generated via the one-hit model.The phenotypes of Arabidopsis and Medicago ta-siRNA mutants, while strikingly different, likewise result from misexpression of the tasiR-ARF target ARF3.Instead, we propose that divergence in the gene networks downstream of the ARF3 transcription factors or the spatiotemporal pattern during leaf development in which these proteins act constitute key factors underlying the distinct contributions of the ta-siRNA pathway to development in maize, Arabidopsis, and possibly other plant species as well.

View Article: PubMed Central - PubMed

Affiliation: Cold Spring Harbor Laboratory, Cold Spring Harbor, New York, United States of America.

ABSTRACT
Maize leafbladeless1 (lbl1) encodes a key component in the trans-acting short-interfering RNA (ta-siRNA) biogenesis pathway. Correlated with a great diversity in ta-siRNAs and the targets they regulate, the phenotypes conditioned by mutants perturbing this small RNA pathway vary extensively across species. Mutations in lbl1 result in severe developmental defects, giving rise to plants with radial, abaxialized leaves. To investigate the basis for this phenotype, we compared the small RNA content between wild-type and lbl1 seedling apices. We show that LBL1 affects the accumulation of small RNAs in all major classes, and reveal unexpected crosstalk between ta-siRNA biogenesis and other small RNA pathways regulating transposons. Interestingly, in contrast to data from other plant species, we found no evidence for the existence of phased siRNAs generated via the one-hit model. Our analysis identified nine TAS loci, all belonging to the conserved TAS3 family. Information from RNA deep sequencing and PARE analyses identified the tasiR-ARFs as the major functional ta-siRNAs in the maize vegetative apex where they regulate expression of AUXIN RESPONSE FACTOR3 (ARF3) homologs. Plants expressing a tasiR-ARF insensitive arf3a transgene recapitulate the phenotype of lbl1, providing direct evidence that deregulation of ARF3 transcription factors underlies the developmental defects of maize ta-siRNA biogenesis mutants. The phenotypes of Arabidopsis and Medicago ta-siRNA mutants, while strikingly different, likewise result from misexpression of the tasiR-ARF target ARF3. Our data indicate that diversity in TAS pathways and their targets cannot fully account for the phenotypic differences conditioned by ta-siRNA biogenesis mutants across plant species. Instead, we propose that divergence in the gene networks downstream of the ARF3 transcription factors or the spatiotemporal pattern during leaf development in which these proteins act constitute key factors underlying the distinct contributions of the ta-siRNA pathway to development in maize, Arabidopsis, and possibly other plant species as well.

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lbl1 affects small RNA biogenesis at select loci.(A) Compared to wild-type, lbl1-rgd1 mutant seedlings show a reduced stature and develop radial, abaxialized leaves. (B) lbl1 has a relatively subtle effect on the overall small RNA population; in three independent biological replicates 79, 172, and 209 loci generate 21-, 22-, or 24-nt small RNAs, respectively, that are significantly changed (q-value<0.05) at least 2-fold between wild-type and lbl1. Grey dots mark the relative normalized abundance of small RNAs not significantly changed between wild-type and lbl1. Colored dots correspond to significantly changed small RNAs: red, 21-nt; blue, 22-nt; green, 24-nt. (C) Low copy genic regions accumulating significantly fewer 21-nt (left), 22-nt (middle) and/or 24-nt (right) siRNAs in lbl1. Genes marked in bold and underlined show reduced levels for all three siRNA classes. Genes marked in bold show reduced 21- and 22-nt siRNAs. Note that the abundance of 21-nt siRNAs at these loci is generally substantially higher. Values are reported on a log scale and represent the mean normalized read counts (RPM) and standard deviation across three independent biological replicates. * adjusted p-value<0.05; ** adjusted p-value<0.01.
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pgen-1004826-g001: lbl1 affects small RNA biogenesis at select loci.(A) Compared to wild-type, lbl1-rgd1 mutant seedlings show a reduced stature and develop radial, abaxialized leaves. (B) lbl1 has a relatively subtle effect on the overall small RNA population; in three independent biological replicates 79, 172, and 209 loci generate 21-, 22-, or 24-nt small RNAs, respectively, that are significantly changed (q-value<0.05) at least 2-fold between wild-type and lbl1. Grey dots mark the relative normalized abundance of small RNAs not significantly changed between wild-type and lbl1. Colored dots correspond to significantly changed small RNAs: red, 21-nt; blue, 22-nt; green, 24-nt. (C) Low copy genic regions accumulating significantly fewer 21-nt (left), 22-nt (middle) and/or 24-nt (right) siRNAs in lbl1. Genes marked in bold and underlined show reduced levels for all three siRNA classes. Genes marked in bold show reduced 21- and 22-nt siRNAs. Note that the abundance of 21-nt siRNAs at these loci is generally substantially higher. Values are reported on a log scale and represent the mean normalized read counts (RPM) and standard deviation across three independent biological replicates. * adjusted p-value<0.05; ** adjusted p-value<0.01.

Mentions: In contrast to Arabidopsis, loss of AGO7 activity in Medicago results in the formation of highly lobed leaves [14], and mutants defective for ta-siRNA biogenesis components in rice and tomato exhibit severe defects in meristem maintenance, mediolateral blade expansion, and adaxial-abaxial leaf polarity [15],[16]. Likewise, mutations in maize lbl1 and ragged seedling2 (rgd2), which encode the orthologs of SGS3 and AGO7, respectively, have severe effects on meristem function and leaf development [17]–[18]. lbl1 mutants, in particular, develop radial, fully abaxialized leaves (Fig. 1A).


Genome-wide analysis of leafbladeless1-regulated and phased small RNAs underscores the importance of the TAS3 ta-siRNA pathway to maize development.

Dotto MC, Petsch KA, Aukerman MJ, Beatty M, Hammell M, Timmermans MC - PLoS Genet. (2014)

lbl1 affects small RNA biogenesis at select loci.(A) Compared to wild-type, lbl1-rgd1 mutant seedlings show a reduced stature and develop radial, abaxialized leaves. (B) lbl1 has a relatively subtle effect on the overall small RNA population; in three independent biological replicates 79, 172, and 209 loci generate 21-, 22-, or 24-nt small RNAs, respectively, that are significantly changed (q-value<0.05) at least 2-fold between wild-type and lbl1. Grey dots mark the relative normalized abundance of small RNAs not significantly changed between wild-type and lbl1. Colored dots correspond to significantly changed small RNAs: red, 21-nt; blue, 22-nt; green, 24-nt. (C) Low copy genic regions accumulating significantly fewer 21-nt (left), 22-nt (middle) and/or 24-nt (right) siRNAs in lbl1. Genes marked in bold and underlined show reduced levels for all three siRNA classes. Genes marked in bold show reduced 21- and 22-nt siRNAs. Note that the abundance of 21-nt siRNAs at these loci is generally substantially higher. Values are reported on a log scale and represent the mean normalized read counts (RPM) and standard deviation across three independent biological replicates. * adjusted p-value<0.05; ** adjusted p-value<0.01.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4263373&req=5

pgen-1004826-g001: lbl1 affects small RNA biogenesis at select loci.(A) Compared to wild-type, lbl1-rgd1 mutant seedlings show a reduced stature and develop radial, abaxialized leaves. (B) lbl1 has a relatively subtle effect on the overall small RNA population; in three independent biological replicates 79, 172, and 209 loci generate 21-, 22-, or 24-nt small RNAs, respectively, that are significantly changed (q-value<0.05) at least 2-fold between wild-type and lbl1. Grey dots mark the relative normalized abundance of small RNAs not significantly changed between wild-type and lbl1. Colored dots correspond to significantly changed small RNAs: red, 21-nt; blue, 22-nt; green, 24-nt. (C) Low copy genic regions accumulating significantly fewer 21-nt (left), 22-nt (middle) and/or 24-nt (right) siRNAs in lbl1. Genes marked in bold and underlined show reduced levels for all three siRNA classes. Genes marked in bold show reduced 21- and 22-nt siRNAs. Note that the abundance of 21-nt siRNAs at these loci is generally substantially higher. Values are reported on a log scale and represent the mean normalized read counts (RPM) and standard deviation across three independent biological replicates. * adjusted p-value<0.05; ** adjusted p-value<0.01.
Mentions: In contrast to Arabidopsis, loss of AGO7 activity in Medicago results in the formation of highly lobed leaves [14], and mutants defective for ta-siRNA biogenesis components in rice and tomato exhibit severe defects in meristem maintenance, mediolateral blade expansion, and adaxial-abaxial leaf polarity [15],[16]. Likewise, mutations in maize lbl1 and ragged seedling2 (rgd2), which encode the orthologs of SGS3 and AGO7, respectively, have severe effects on meristem function and leaf development [17]–[18]. lbl1 mutants, in particular, develop radial, fully abaxialized leaves (Fig. 1A).

Bottom Line: Interestingly, in contrast to data from other plant species, we found no evidence for the existence of phased siRNAs generated via the one-hit model.The phenotypes of Arabidopsis and Medicago ta-siRNA mutants, while strikingly different, likewise result from misexpression of the tasiR-ARF target ARF3.Instead, we propose that divergence in the gene networks downstream of the ARF3 transcription factors or the spatiotemporal pattern during leaf development in which these proteins act constitute key factors underlying the distinct contributions of the ta-siRNA pathway to development in maize, Arabidopsis, and possibly other plant species as well.

View Article: PubMed Central - PubMed

Affiliation: Cold Spring Harbor Laboratory, Cold Spring Harbor, New York, United States of America.

ABSTRACT
Maize leafbladeless1 (lbl1) encodes a key component in the trans-acting short-interfering RNA (ta-siRNA) biogenesis pathway. Correlated with a great diversity in ta-siRNAs and the targets they regulate, the phenotypes conditioned by mutants perturbing this small RNA pathway vary extensively across species. Mutations in lbl1 result in severe developmental defects, giving rise to plants with radial, abaxialized leaves. To investigate the basis for this phenotype, we compared the small RNA content between wild-type and lbl1 seedling apices. We show that LBL1 affects the accumulation of small RNAs in all major classes, and reveal unexpected crosstalk between ta-siRNA biogenesis and other small RNA pathways regulating transposons. Interestingly, in contrast to data from other plant species, we found no evidence for the existence of phased siRNAs generated via the one-hit model. Our analysis identified nine TAS loci, all belonging to the conserved TAS3 family. Information from RNA deep sequencing and PARE analyses identified the tasiR-ARFs as the major functional ta-siRNAs in the maize vegetative apex where they regulate expression of AUXIN RESPONSE FACTOR3 (ARF3) homologs. Plants expressing a tasiR-ARF insensitive arf3a transgene recapitulate the phenotype of lbl1, providing direct evidence that deregulation of ARF3 transcription factors underlies the developmental defects of maize ta-siRNA biogenesis mutants. The phenotypes of Arabidopsis and Medicago ta-siRNA mutants, while strikingly different, likewise result from misexpression of the tasiR-ARF target ARF3. Our data indicate that diversity in TAS pathways and their targets cannot fully account for the phenotypic differences conditioned by ta-siRNA biogenesis mutants across plant species. Instead, we propose that divergence in the gene networks downstream of the ARF3 transcription factors or the spatiotemporal pattern during leaf development in which these proteins act constitute key factors underlying the distinct contributions of the ta-siRNA pathway to development in maize, Arabidopsis, and possibly other plant species as well.

Show MeSH
Related in: MedlinePlus