Identification of miRNAs with potential roles in regulation of anther development and male-sterility in 7B-1 male-sterile tomato mutant

Abstract

BackgroundThe 7B-1 tomato line (Solanum lycopersicum cv. Rutgers) is a photoperiod-sensitive male-sterile mutant, with potential application in hybrid seed production. Small RNAs (sRNAs) in tomato have been mainly characterized in fruit development and ripening, but none have been studied with respect to flower development and regulation of male-sterility. Using sRNA sequencing, we identified miRNAs that are potentially involved in anther development and regulation of male-sterility in 7B-1 mutant.ResultsTwo sRNA libraries from 7B-1 and wild type (WT) anthers were sequenced and thirty two families of known miRNAs and 23 new miRNAs were identified in both libraries. MiR390, miR166, miR159 were up-regulated and miR530, miR167, miR164, miR396, miR168, miR393, miR8006 and two new miRNAs, miR#W and miR#M were down-regulated in 7B-1 anthers. Ta-siRNAs were not differentially expressed and likely not associated with 7B-1 male-sterility. miRNA targets with potential roles in anther development were validated using 5′-RACE. QPCR analysis showed differential expression of miRNA/target pairs of interest in anthers and stem of 7B-1, suggesting that they may regulate different biological processes in these tissues. Expression level of most miRNA/target pairs showed negative correlation, except for few. In situ hybridization showed predominant expression of miR159, GAMYBL1, PMEI and cystatin in tapetum, tetrads and microspores.ConclusionOverall, we identified miRNAs with potential roles in anther development and regulation of male-sterility in 7B-1. A number of new miRNAs were also identified from tomato for the first time. Our data could be used as a benchmark for future studies of the molecular mechanisms of male-sterility in other crops.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-2077-0) contains supplementary material, which is available to authorized users.

Distinct transcriptional and processing regulations control miR167a level in tomato during stress

“…(1) Its level in leaf tissues was increased with the progression of development from plumule to the formation of the first true leaf, (2) it accumulated more in light grown seedlings compared to the seedlings grown in dark, and (3) its level was dependent on the specific stress applied to the plant. The expression pattern of targets of miR167a 14,15 was also in agreement with the corresponding level of the miRNA in some of the conditions tested (Fig. 2B-E).…”

“…But, how all these processes contribute in determining the steady state level of a mature miRNA in the plant cell remains elusive. In this regard, miR167a in tomato plant served as a model system to study the complex regulation, as the miRNA has definite role in plant developmental processes [13][14][15] with potential implications in stress physiology. 12 Thus, here we have attempted to untangle the various regulatory modules of miR167a to gain insight to the mechanism of regulation of miR167a expression and/or de novo biogenesis on demand.…”

“…10 miR167 is an evolutionarily conserved miRNA 11 which performs important regulatory role in flower development, adventitious rooting and nodulation along with probable roles in biotic and abiotic stress response. [12][13][14][15] Available literature 13,15 depicts that miR167 negatively regulates ARF6/8 and IAA-Ala Resistant 3. Recent study by 16 showed that blue light augmented miR167 expression, resulting inhibition of ARFs and modulation of photomorphogenesis in Arabidopsis.…”

Comprehensive processing of high-throughput small RNA sequencing data including quality checking, normalization, and differential expression analysis using the UEA sRNA Workbench

“…The percentage of genome-matching reads is calculated for both redundant and non-redundant sequences and across size classes. Selected annotations for which similar checks are performed include miRNAs, other ncRNAs (such as tRNAs, rRNAs, or snoRNAs), protein-coding genes, and repeat/transposable elements depending on available annotation information (Xu et al 2014;Omidvar et al 2015).…”

“…These RNA fragments are excised by Dicer/Dicer-like proteins from double-stranded RNA precursors deriving either from single stranded RNAs with a hairpin-like secondary structure, the miRNAs (Zhu et al 2013), or long double-stranded RNA created by a polymerase, the siRNAs (Chen 2012). The sRNAs target and subsequently silence genes and thus play an important role in gene regulation (Lippman and Martienssen 2004;Omidvar et al 2015), defense against pathogens (Szittya et al 2010;Donaszi-Ivanov et al 2013) and general maintenance of the genome (Molnar et al 2007;Mohorianu et al 2011).…”