The bHLH142 Transcription Factor Coordinates with TDR1 to Modulate the Expression of EAT1 and Regulate Pollen Development in Rice    

Abstract

Male sterility plays an important role in F1 hybrid seed production. We identified a male-sterile rice (Oryza sativa) mutant with impaired pollen development and a single T-DNA insertion in the transcription factor gene bHLH142. Knockout mutants of bHLH142 exhibited retarded meiosis and defects in tapetal programmed cell death. RT-PCR and in situ hybridization analyses showed that bHLH142 is specifically expressed in the anther, in the tapetum, and in meiocytes during early meiosis. Three basic helix-loop-helix transcription factors, UDT1 (bHLH164), TDR1 (bHLH5), and EAT1/DTD1 (bHLH141) are known to function in rice pollen development. bHLH142 acts downstream of UDT1 and GAMYB but upstream of TDR1 and EAT1 in pollen development. In vivo and in vitro assays demonstrated that bHLH142 and TDR1 proteins interact. Transient promoter assays demonstrated that regulation of the EAT1 promoter requires bHLH142 and TDR1. Consistent with these results, 3D protein structure modeling predicted that bHLH142 and TDR1 form a heterodimer to bind to the EAT1 promoter. EAT1 positively regulates the expression of AP37 and AP25, which induce tapetal programmed cell death. Thus, in this study, we identified bHLH142 as having a pivotal role in tapetal programmed cell death and pollen development.

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

“…MS23 interacts with bHLH51 but not bHLH122 (Fig. S14) paralleling its counterpart in rice (Fu et al, 2014;Ko et al, 2014). Additionally, MS32 only forms strong dimers with bHLH122 in maize whereas bHLH51 forms both homodimers and strong dimers with bHLH122.…”

“…Mutants of tip2, the rice ortholog of Ms23, have sporadic, periclinal divisions in the pre-tapetal layer but never form a complete bilayer as in ms23-ref (Fu et al, 2014;Ko et al, 2014). Furthermore, in situ hybridization reveals that transcript localizations are distinct in these grasses.…”

“…In Arabidopsis, the ortholog of Ms32 (DYT1) is also upstream of bHLH10, bHLH89, bHLH91 and AMS. Interestingly, Ms32, UDT1 and DYT1 are all upregulated in the mutants corresponding to ms23-ref (rice tip2 and Arabidopsis bhlh10/bhlh89/bhlh91) (Fu et al, 2014;Ko et al, 2014;Zhu et al, 2015).…”

“…S10). By contrast, rice EAT1 positively regulates the expression of TIP2 (Ko et al, 2014). Arabidopsis AMS is regulated by bHLH10, bHLH89 and bHLH91.…”

“…Mutant analysis demonstrates that several tapetal bHLHs are crucial for (1) pre-meiotic to meiotic stages: Arabidopsis DYSFUNCTIONAL TAPETUM1 (DYT1) Feng et al, 2012), its rice homolog UNDEVELOPED TAPETUM1 (UDT1) (Jung et al, 2005), and the maize homolog, Male Sterility32 (Ms32) (Moon et al, 2013); rice ETERNAL TAPETUM1 (EAT1 -OS04G0599300, also named DELAYED TAPETUM DEGENERATION or DTD) (Niu et al, 2013;Ji et al, 2013); rice TDR INTERACTING PARTNER2 (TIP2 -OS01T0293100; also named bHLH142) (Fu et al, 2014;Ko et al, 2014); Arabidopsis bHLH10, bHLH89 and bHLH91 (Zhu et al, 2015); as well as for (2) meiotic to post-meiotic stages: Arabidopsis ABORTED MICROSPORES (AMS) (Sorensen et al, 2003;Xu et al, 2010Xu et al, , 2014 and its rice homolog TAPETUM DEGENERATION RETARDATION (TDR) (Li et al, 2006;Zhang et al, 2008).…”