Transcriptome Analysis of Triple Mutant for OsMADS62, OsMADS63, and OsMADS68 Reveals the Downstream Regulatory Mechanism for Pollen Germination in Rice (Oryza sativa)

Abstract

The MADS (MCM1-AGAMOUS-DEFFICIENS-SRF) gene family has a preserved domain called MADS-box that regulates downstream gene expression as a transcriptional factor. Reports have revealed three MADS genes in rice, OsMADS62, OsMADS63, and OsMADS68, which exhibits preferential expression in mature rice pollen grains. To better understand the transcriptional regulation of pollen germination and tube growth in rice, we generated the loss-of-function homozygous mutant of these three OsMADS genes using the CRISPR-Cas9 (clustered regularly interspaced short palindromic repeats-CRISPR associated protein 9) system in wild-type backgrounds. Results showed that the triple knockout (KO) mutant showed a complete sterile phenotype without pollen germination. Next, to determine downstream candidate genes that are transcriptionally regulated by the three OsMADS genes during pollen development, we proceeded with RNA-seq analysis by sampling the mature anther of the mutant and wild-type. Two hundred and seventy-four upregulated and 658 downregulated genes with preferential expressions in the anthers were selected. Furthermore, downregulated genes possessed cell wall modification, clathrin coat assembly, and cellular cell wall organization features. We also selected downregulated genes predicted to be directly regulated by three OsMADS genes through the analyses for promoter sequences. Thus, this study provides a molecular background for understanding pollen germination and tube growth mediated by OsMADS62, OsMADS63, and OsMADS68 with mature pollen preferred expression.

Genome-Wide Identification of MADS-Box Family Genes in Safflower (Carthamus tinctorius L.) and Functional Analysis of CtMADS24 during Flowering

“…In recent years, with the accomplishment of various whole genome sequencing projects, the study of related gene families is rapidly progressing. Many MADS-box genes have been identified in various plant species including Arabidopsis [ 21 ], rice [ 39 ], sunflower [ 40 ], bamboo [ 41 ], pear [ 42 ], wheat [ 43 ], strawberry [ 44 ], and many other plants. These findings laid the groundwork for studying the evolution of MADS-box genes.…”

“…The discovery of genes related to flower development and flowering transition remains a major focus in plant research. Studies have shown that MADS-box genes belonging to the type I clade are mainly involved in the development of the female gametophyte, the embryo, and seeds, while plant type II MADS-box genes have specific roles in flower organ development, especially in specifying floral organ identity [ 39 , 48 ]. In most plants, different subfamily members often have different expression patterns [ 45 , 49 ].…”

Transcription Factors and Their Regulatory Roles in the Male Gametophyte Development of Flowering Plants

“…How the expression of these genes is coordinated for the sequential male gametophyte development has not been well investigated. Analysis of transcriptomic data identified a set of TFs that are specifically or preferentially expressed during male gametophyte development [ 22 , 51 , 52 , 53 ]. In the last decades, a set of reverse genetic screens and forward genetic strategies have identified a batch of TFs with specific expressions and regulatory roles in male gametophyte development, demonstrating the undoubted implementation of TFs in the pollen ontogenesis of flowering plants.…”

Sucrose preferentially promotes expression of OsWRKY7 and OsPR10a to enhance defense response to blast fungus in rice

“…All the mapped genes were downloaded and filtered based on the following criteria: hyper p-value < 0.05 and query numbers > 10. Additional significance of the GO terms was denoted by a fold enrichment value > 1.5, which was obtained by dividing the query number by the query expectation value (Hong et al, 2020;Kim et al, 2021). The biotic stress category was obtained via the MapMan analysis toolkit 3.5.1 R2 (Thimm et al, 2004).…”