Intragenic suppressors unravel the role of the SCREAM ACT-like domain for bHLH partner selectivity in stomatal development.

Hyemin Seo, Krishna Mohan Sepuru, Aarthi Putarjunan, Lyndsey Aguirre, Benjamin A Burrows, Keiko U Torii

Proceedings of the National Academy of Sciences of the United States of America 2022 Mar 01

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Multicellular organisms develop specialized cell types to achieve complex functions of tissues and organs. The basic helix-loop-helix (bHLH) proteins act as master regulatory transcription factors of such specialized cell types. Plant stomata are cellular valves in the aerial epidermis for efficient gas exchange and water control. Stomatal differentiation is governed by sequential actions of three lineage-specific bHLH proteins, SPEECHLESS (SPCH), MUTE, and FAMA, specifying initiation and proliferation, commitment, and terminal differentiation, respectively. A broadly expressed bHLH, SCREAM (SCRM), heterodimerizes with SPCH/MUTE/FAMA and drives stomatal differentiation via switching its partners. Yet nothing is known about its heterodimerization properties or partner preference. Here, we report the role of the SCRM C-terminal ACT-like (ACTL) domain for heterodimerization selectivity. Our intragenic suppressor screen of a dominant scrm-D mutant identified the ACTL domain as a mutation hotspot. Removal of this domain or loss of its structural integrity abolishes heterodimerization with MUTE, but not with SPCH or FAMA, and selectively abrogates the MUTE direct target gene expression. Consequently, the scrm-D ACTL mutants confer massive clusters of arrested stomatal precursor cells that cannot commit to differentiation when redundancy is removed. Structural and biophysical studies further show that SPCH, MUTE, and FAMA also possess the C-terminal ACTL domain, and that ACTL•ACTL heterodimerization is sufficient for partner selectivity. Our work elucidates a role for the SCRM ACTL domain in the MUTE-governed proliferation-differentiation switch and suggests mechanistic insight into the biological function of the ACTL domain, a module uniquely associated with plant bHLH proteins, as a heterodimeric partner selectivity interface.

Citation

Hyemin Seo, Krishna Mohan Sepuru, Aarthi Putarjunan, Lyndsey Aguirre, Benjamin A Burrows, Keiko U Torii. Intragenic suppressors unravel the role of the SCREAM ACT-like domain for bHLH partner selectivity in stomatal development. Proceedings of the National Academy of Sciences of the United States of America. 2022 Mar 01;119(9)