Global transcriptome analysis of two ameiotic1 alleles in maize anthers: defining steps in meiotic entry and progression through prophase I

Abstract

BackgroundDevelopmental cues to start meiosis occur late in plants. Ameiotic1 (Am1) encodes a plant-specific nuclear protein (AM1) required for meiotic entry and progression through early prophase I. Pollen mother cells (PMCs) remain mitotic in most am1 mutants including am1-489, while am1-praI permits meiotic entry but PMCs arrest at the leptotene/zygotene (L/Z) transition, defining the roles of AM1 protein in two distinct steps of meiosis. To gain more insights into the roles of AM1 in the transcriptional pre-meiotic and meiotic programs, we report here an in depth analysis of gene expression alterations in carefully staged anthers at 1 mm (meiotic entry) and 1.5 mm (L/Z) caused by each of these am1 alleles.Results1.0 mm and 1.5 mm anthers of am1-489 and am1-praI were profiled in comparison to fertile siblings on Agilent® 4 × 44 K microarrays. Both am1-489 and am1-praI anthers are cytologically normal at 1.0 mm and show moderate transcriptome alterations. At the 1.5-mm stage both mutants are aberrant cytologically, and show more drastic transcriptome changes. There are substantially more absolute On/Off and twice as many differentially expressed genes (sterile versus fertile) in am1-489 than in am1-praI. At 1.5 mm a total of 4,418 genes are up- or down-regulated in either am1-489 or am1-praI anthers. These are predominantly stage-specific transcripts. Many putative meiosis-related genes were found among them including a small subset of allele-specific, mis-regulated genes specific to the PMCs. Nearly 60% of transcriptome changes in the set of transcripts mis-regulated in both mutants (N = 530) are enriched in PMCs, and only 1% are enriched in the tapetal cell transcriptome. All array data reported herein will be deposited and accessible at MaizeGDB http://www.maizegdb.org/.ConclusionsOur analysis of anther transcriptome modulations by two distinct am1 alleles, am1-489 and am1-praI, redefines the role of AM1 as a modulator of expression of a subset of meiotic genes, important for meiotic progression and provided stage-specific insights into the genetic networks associated with meiotic entry and early prophase I progression.

The transcriptome landscape of early maize meiosis

“…Twenty of our meiosis-candidate genes were contained in the list, most notably Am1 and Phs1 which have already been shown in maize to be involved in meiotic recombination and whose loss results in male sterility [16,32,36]. …”

Spatiotemporally dynamic, cell-type–dependent premeiotic and meiotic phasiRNAs in maize anthers

“…Together with the conclusions from analysis of ocl4, these data suggest that in both premeiotic and meiotic phasiRNA pathways, the production of the miRNA triggers and the PHAS precursors are separately regulated. To test whether normal meiocytes are required for phasiRNAs production, we sequenced anther sRNAs from ameiotic1 (am1), in which somatic lobe cells are normal but PMCs or meiocytes are defective (25). Two am1 alleles, am1-489 (PMCs conduct mitosis instead of meiosis) and am1-praI (meiocytes arrest in prophase I), have premeiotic and meiotic phasiRNAs (Fig.…”

MS23, a master basic helix-loop-helix factor, regulates the specification and development of tapetum in maize

“…The periclinal divisions of both PPCs and SPCs appeared normal in ms23-ref anthers, in terms of initiation timing and completion of a four-layer anther wall. In fertile anthers, the pre-meiotic archesporial cells completed mitotic divisions before Stage 9, then matured into the PMCs; these synthesized DNA and organized the meiotic chromosomes then entered prophase I, progressed through zygotene, and entered meiosis II at Stages 8, 9 and 10, respectively (Nan et al, 2011). During the pre-meiotic and meiotic stages, somatic wall layer cells continued to divide anticlinally to add anther width and length.…”

“…Total maize RNA was isolated using the PureLink RNA Mini Kit (Invitrogen) incorporating TRIzol Reagent (Invitrogen) with an oncolumn DNase I (Qiagen) digestion according to the manufacturer's instructions. Microarray experiments were conducted as previously described (Nan et al, 2011). qRT-PCR was performed as previously described with three biological samples and three technical replicates unless stated otherwise .…”