Genome-Wide Identification of miRNAs and Their Targets Involved in the Developing Internodes under Maize Ears by Responding to Hormone Signaling

<div><p>Internode length is one of the decisive factors affecting plant height (PH) and ear height (EH), which are closely associated with the lodging resistance, biomass and grain yield of maize. miRNAs, currently recognized as important transcriptional/ post-transcriptional regulators, play an essential role in plant growth and development. However, their roles in developing internodes under maize ears remain unclear. To identify the roles of miRNAs and their targets in the development of internodes under maize ears, six miRNA and two degradome libraries were constructed using the 7^th, 8^th and 9^th internodes of two inbred lines, ‘Xun928’ and ‘Xun9058’, which had significantly different internode lengths. A total of 45 and 54 miRNAs showed significant changes for each pairwise comparison among the 7^th, 8^th and 9^th internodes of ‘Xun9058’ and ‘Xun928’, respectively. The expression of 31 miRNAs showed significant changes were common to the corresponding comparison groups of the 7^th, 8^th and 9^th internodes of ‘Xun9058’ and ‘Xun928’. For the corresponding internodes of ‘Xun9058’ and ‘Xun928’, compared with the expression of miRNAs in the 7^th, 8^th and 9^th internodes of ‘Xun928’, the numbers of up-regulated and down-regulated miRNAs were 11 and 36 in the 7^th internode, 9 and 45 in the 8^th internode, and 9 and 25 in the 9^th internode of ‘Xun9058’, respectively. Moreover, 10 miRNA families containing 45 members showed significant changes at least in two internodes of ‘Xun928’ by comparing with the corresponding internodes of ‘Xun9058’. Based on the sequencing data, 20 miRNAs related to hormone signaling among the candidates, belonging to five conserved miRNA families, were selected for expression profiling using quantitative reverse-transcription polymerase chain reaction (qRT-PCR). The five miRNA families, zma-miR160, zma-miR167, zma-miR164, zma-miR169 and zma-miR393, targeted the genes encoding auxin response factor, N-acetylcysteine domain containing protein, nuclear transcription factor Y and auxin signaling F-BOX 2 through degradome sequencing. The miRNAs might regulate their targets to respond to hormone signaling, thereby regulating the internode elongation and development under maize ear. These results provide valuable reference for understanding the possible regulation mechanism of the ILs under the ear.</p></div

Conserved miRNA family members in the six maize internode libraries.

<p>miR166 consisted of 14 members, miR156 and miR167 had 12 and 10 members, respectively, and miR162, miR529, miR827 and miR1432 had only one member.</p

Expression levels of maize internode-associated miRNAs as determined by RT-PCR.

<p>18S rRNA was used as the reference gene. Three biological replicates and three technical replicates were performed. Data represent the mean values ± SD of three replicates.</p

Average IL (cm) of the 7^th to the 9^th internodes in ‘Xun928’ and ‘Xun9058’.

<p>Average IL (cm) of the 7^th to the 9^th internodes in ‘Xun928’ and ‘Xun9058’.</p

Length distribution of small RNAs obtained from the six maize internode libraries.

<p>The 24 nt was the most abundant size of the small RNAs, accounting for ~60% at each internode.</p