Pollen Number and Ribosome Gene Expression Altered in a Genome-Editing Mutant of REDUCED POLLEN NUMBER1 Gene

Abstract

The number of pollen grains varies within and between species. However, little is known about the molecular basis of this quantitative trait, in contrast with the many studies available on cell differentiation in the stamen. Recently, the first gene responsible for pollen number variation, REDUCED POLLEN NUMBER1 (RDP1), was isolated by genome-wide association studies of Arabidopsis thaliana and exhibited the signature of natural selection. This gene encodes a homolog of yeast Mrt4 (mRNA turnover4), which is an assembly factor of the large ribosomal subunit. However, no further data were available to link ribosome function to pollen development. Here, we characterized the RDP1 gene using the standard A. thaliana accession Col-0. The frameshift mutant, rdp1-3 generated by CRISPR/Cas9 revealed the pleiotropic effect of RDP1 in flowering, thus demonstrating that this gene is required for a broad range of processes other than pollen development. We found that the natural Col-0 allele conferred a reduced pollen number against the Bor-4 allele, as assessed using the quantitative complementation test, which is more sensitive than transgenic experiments. Together with a historical recombination event in Col-0, which was identified by sequence alignment, these results suggest that the coding sequence of RDP1 is the candidate region responsible for the natural phenotypic variation. To elucidate the biological processes in which RDP1 is involved, we conducted a transcriptome analysis. We found that genes responsible for ribosomal large subunit assembly/biogenesis were enriched among the differentially regulated genes, which supported the hypothesis that ribosome biogenesis is disturbed in the rdp1-3 mutant. Among the pollen-development genes, three key genes encoding basic helix-loop-helix (bHLH) transcription factors (ABORTED MICROSPORES (AMS), bHLH010, and bHLH089), as well as direct downstream genes of AMS, were downregulated in the rdp1-3 mutant. In summary, our results suggest a specialized function of ribosomes in pollen development through RDP1, which harbors natural variants under selection.

Genotyping by Sequencing for SNP-Based Linkage Analysis and the Development of KASPar Markers for Male Sterility and Polyembryony in Citrus

“…Pollen grain number in angiosperms is a key reproductive trait that has been studied extensively for decades. Despite its agricultural and evolutionary importance, the genetic basis of the pollen grain number has remained elusive, primarily due to its quantitative nature [ 72 ]. The information generated from gene annotations allows us to focus our efforts on 19 genes related to male sterility over the 67 genes annotated in the genomic region identified by the QTL analysis.…”

Genome-wide association study highlights escape from aphids by delayed growth inArabidopsis thaliana

“…In our study, the other growth-related genes or other mutations of AT3G13882 might have reduced the aphid abundance. Because transgenic approaches may not be effective to identify mutation sites affecting quantitative traits, further experimental tests, such as quantitative complementation [16,40], will be needed to identify natural causal variants that alter aphid abundance through delayed growth.…”

“…While ribosomal genes have long been considered housekeeping genes of the protein synthesis machinery, mutants of ribosome-related genes exhibit a wide variety of growth and reproductive phenotypes. For example, previous studies reported the reduction of leaf cell number [38], reduced root length [39], and the reduction of pollen number [16,40] regarding ribosomal gene mutations. In this context, our findings from the ribosomal gene AT3G13882 add insights into biotic interactions besides growth deterioration.…”

“…Multiple alleles on the AT3G13882 gene are also needed to provide strong evidence of those phenotypes. In the studies of a ribosomal gene REDUCED POLLEN NUMBER1 ( RDP1 ), null mutants showed a pleiotropic effect on plant growth and pollen number in A. thaliana [16,40]. Natural alleles of RDP1 could alleviate pleiotropic growth defects [16].…”

mRNA Turnover Protein 4 Is Vital for Fungal Pathogenicity and Response to Oxidative Stress in Sclerotinia sclerotiorum

“…In addition, MRT4 plays an important role in cell tolerance in yeast [ 19 ]. The loss of At RDP1 (the homolog of MRT4) could decrease the amount of pollen in Arabidopsis thaliana and inhibit its development, possibly due to ribosome specialization [ 20 ]. It was shown that the subcellular localization of human MRT4 is regulated by the C-terminal region under stress [ 21 ].…”