MYB81, a microspore‐specific GAMYB transcription factor, promotes pollen mitosis I and cell lineage formation in Arabidopsis

Abstract

Sexual reproduction in flowering plants relies on the production of haploid gametophytes that consist of germline and supporting cells. During male gametophyte development, the asymmetric mitotic division of an undetermined unicellular microspore segregates these two cell lineages. To explore genetic regulation underlying this process, we screened for pollen cell patterning mutants and isolated the heterozygous myb81-1 mutant that sheds~50% abnormal pollen. Typically, myb81-1 microspores fail to undergo pollen mitosis I (PMI) and arrest at polarized stage with a single central vacuole. Although most myb81-1 microspores degenerate without division, a small fraction divides at later stages and fails to acquire correct cell fates. The myb81-1 allele is transmitted normally through the female, but rarely through pollen. We show that myb81-1 phenotypes result from impaired function of the GAMYB transcription factor MYB81. The MYB81 promoter shows microspore-specific activity and a MYB81-RFP fusion protein is only expressed in a narrow window prior to PMI. Ectopic expression of MYB81 driven by various promoters can severely impair vegetative or reproductive development, reflecting the strict microspore-specific control of MYB81. Our data demonstrate that MYB81 has a key role in the developmental progression of microspores, enabling formation of the two male cell lineages that are essential for sexual reproduction in Arabidopsis.

Analysis of sticky generative cell mutants reveals that suppression of callose deposition in the generative cell is necessary for generative cell internalization and differentiation in Arabidopsis

“…To examine whether the sgc mutation also affected cell fate specification, we generated wild‐type and sgc‐1 mutant plants expressing both vegetative cell‐ and germline‐specific markers using a doubly homozygous marker line harboring proLAT52‐H2B‐GFP and proMGH3‐H2B‐mRFP1 vectors, generated previously by Oh et al . (2020). The promoters of late anther tomato 52 ( LAT52 ) and the male gamete‐specific histone H3.3 ( MGH3 ) genes are well known to drive vegetative cell‐specific (Twell, 1992) and germline‐specific (Okada et al ., 2005) gene expression, respectively.…”

“…2016). For colocalization tests, proSCP‐H2B‐GFP6 and proSCP‐VHAA‐mRFP1 vectors were used to label the nucleus and tonoplast during pollen development (Oh et al ., 2010b; Oh et al ., 2020). In addition, fragments of STtmd, MLS and mCherryHDEL were amplified with forward and reverse primers containing two different restriction enzyme sites (Table S3); these were then subcloned to produce proUBQ14‐STtmd‐GFP, proSGC‐STtmd‐GFP, proUBQ14‐MLS‐GFP, proSGC‐MLS‐GFP and proSGC‐mCherryHDEL vectors.…”

“…Examples include: Arabidopsis and Nicotiana tabacum (tobacco) orthologs, GEM1 / MOR1 and TMBP200 , of the evolutionarily conserved MAP215 microtubule‐associated protein family (Park et al ., 1998; Twell et al ., 2002; Oh et al ., 2010a); two γ‐tubulin isoforms, TUBG1 and TUBG2 (Pastuglia et al ., 2006); a γ‐TuRC‐anchoring factor known as NEDD1 (Zeng et al ., 2009); multiple subunits of the AUGMIN complex (Ho et al ., 2011; Hotta et al ., 2012; Oh et al ., 2016); the Arabidopsis ortholog TWO‐IN‐ONE of the FUSED kinase family (Oh et al ., 2005; Oh et al ., 2012); two pairs of kinesin genes, KINESIN‐12A and KINESIN‐12B (Lee et al ., 2007); and HINKEL and TETRASPORE (Oh et al ., 2008). In addition, two microspore‐specific transcription factor genes suggest the presence of genetic networks for pollen mitosis I: SIDECAR POLLEN ( SCP ; also known as LATERAL BOUNDARY DOMAIN 27, LBD27 ), which is involved in the correct timing and division orientation (Chen and McCormick, 1996; Oh et al ., 2010b); and the GAMYB transcription factor MYB81 , which functions in mitotic entry (Oh et al ., 2020).…”

The transcription factors and pathways underpinning male reproductive development in Arabidopsis

“…The larger vegetative cell is responsible for forming the pollen tube, which will deliver the male gametes to the female reproductive structures during fertilization, while the smaller generative cell will eventually divide into two sperm cells through PMII, which occurs prior to pollen maturation. One TF essential for PMI is MYB81, a microspore-specific TF; myb81 microspores are unable to undergo PMI, arresting at the polarized microspore stage ( Oh et al., 2020 ). However, the exact mechanisms through which MYB81 promotes microspore progression into PMI remains unclear.…”

Epigenetic Dynamics and Regulation of Plant Male Reproduction

“…A regulatory cascade AtDYT1-AtTDF1-AtAMS-AtMS188-AtMS1 has been well established in Arabidopsis [ 13 ], and the TF orthologs of the cascade have been proven to play similar roles in rice [ 14 ]. In microgametogenesis, key TFs that are involved in asymmetric division, cell cycle progression, and patterning, such as LBD27 controlling pollen mitosis I (PMI) entry and correct asymmetric division [ 15 ], MYB81 associated with PMI, and daughter cell lineage formation [ 16 ], have been characterized. Readers interested in the genetic regulation of male gametophyte development can refer to available reviews [ 17 , 18 , 19 , 20 ].…”