GORI, encoding the WD40 domain protein, is required for pollen tube germination and elongation in rice

Abstract

Successful delivery of sperm cells to the embryo sac in higher plants is mediated by pollen tube growth. The molecular mechanisms underlying pollen germination and tube growth in crop plants remain rather unclear, although these mechanisms are crucial to plant reproduction and seed formation. By screening pollen-specific gene mutants in rice (Oryza sativa), we identified a T-DNA insertional mutant of Germinating modulator of rice pollen (GORI) that showed a one-to-one segregation ratio for wild type (WT) to heterozygous. GORI encodes a seven-WD40-motif protein that is homologous to JINGUBANG/REN4 in Arabidopsis. GORI is specifically expressed in rice pollen, and its protein is localized in the nucleus, cytosol and plasma membrane. Furthermore, a homozygous mutant, gori-2, created through CRISPR-Cas9 clearly exhibited male sterility with disruption of pollen tube germination and elongation. The germinated pollen tube of gori-2 exhibited decreased actin filaments and altered pectin distribution. Transcriptome analysis revealed that 852 pollen-specific genes were downregulated in gori-2 compared with the WT, and Gene Ontology enrichment analysis indicated that these genes are strongly associated with cell wall modification and clathrin coat assembly. Based on the molecular features of GORI, phenotypical observation of the gori mutant and its interaction with endocytic proteins and Rac GTPase, we propose that GORI plays key roles in forming endo-/exocytosis complexes that could mediate pollen tube growth in rice.

Global Identification of ANTH Genes Involved in Rice Pollen Germination and Functional Characterization of a Key Member, OsANTH3

“…GORI encodes a seven WD40 motif protein that is homologous to REN4 in Arabidopsis. Knock-out of GORI caused 20% pollen germination percentage but germinated pollen has defect on tube elongation as well, causing full male sterility (Kim et al, 2021). REN4 was shown to interact with PICALM9b/EAP1 (Li et al, 2018), similarly to interactions between GORI and OsANTH3.…”

“…In the Y2H screening for the identification of direct protein interactions (Supplementary Figure 7), the C-terminal of OsANTH3 (1096∼end) was found to interact with the PME31 (pectin methylesterase) (Kim Y. et al, 2020) and GORI proteins (Kim et al, 2021), which are both important for rice pollen germination. GORI encodes a seven WD40 motif protein that is homologous to REN4 in Arabidopsis.…”

“…The full length, N-terminal (1∼1,095 bp), and C-terminal (1,096∼end) CDSs of ANTH3 were cloned at the BamHI site of the pGADT7-Rec vector. The full-length, N-terminal, and C-terminal CDSs of GORI were cloned at the EcoRI/BamHI site of the pGBKT7 vector (Kim et al, 2021). The C-terminal (633end) CDS of OsPME31 (Kim Y. et al, 2020) was cloned into the pGBKT7 vector.…”

A Systemic View of Carbohydrate Metabolism in Rice to Facilitate Productivity

“…This expression pattern was further confirmed by qRT-PCR, with samples in six tissues/organs ( Figure 4 b) matching the dissected model. Recently, a defect in the GORI gene changed the distribution of pectin in germinated pollen tubes and eventually led to the male sterile phenotype [ 35 ]. Thus, qRT-PCR was performed to analyze their regulatory roles by GORI in the gori mutant.…”

“…One study reported that glycolysis could regulate pollen tube polarity in Arabidopsis [ 93 ]. Similarly, a recently characterized gene ( GORI ) involved in late pollen development in rice showed its connectivity to carbohydrate metabolism such as less pectin staining in the gori pollen tube [ 35 ]. Interestingly, three genes in cluster H showed significantly reduced expression when GORI was knocked out ( Figure 4 c).…”