The Arabidopsis Histone Chaperone FACT: Role of the HMG-Box Domain of SSRP1

摘要

Histone chaperones play critical roles in regulated structural transitions of chromatin in eukaryotic cells that involve nucleosome disassembly and reassembly. The histone chaperone FACT is a heterodimeric complex consisting in plants and metazoa of SSRP1/SPT16 and is involved in dynamic nucleosome reorganization during various DNA-dependent processes including transcription, replication and repair. The C-terminal HMG-box domain of the SSRP1 subunit mediates interactions with DNA and nucleosomesin vitro, but its relevancein vivois unclear. Here, we demonstrate thatArabidopsis ssrp1–2mutant plants express a C-terminally truncated SSRP1 protein. Although the structure of the truncated HMG-box domain is distinctly disturbed, it still exhibits residual DNA-binding activity, but has lost DNA-bending activity. Sincessrp1–2plants are phenotypically affected but viable, the HMG-box domain may be functionally non-essential. To examine this possibility, SSRP1?HMG completely lacking the HMG-box domain was studied. SSRP1?HMGin vitrodid not bind to DNA and its interactions with nucleosomes were severely reduced. Nevertheless, the protein showed a nuclear mobility and protein interactions similar to SSRP1. Interestingly, expression of SSRP1?HMG is almost as efficient as that of full-length SSRP1 in supporting normal growth and development of the otherwise non-viableArabidopsis ssrp1–1mutant. SSRP1?HMG is structurally similar to the fungal ortholog termed Pob3 that shares clear similarity with SSRP1, but it lacks the C-terminal HMG-box. Therefore, our findings indicate that the HMG-box domain conserved among SSRP1 proteins is not critical inArabidopsis, and thus, the functionality of SSRP1/SPT16 in plants/metazoa and Pob3/Spt16 in fungi is perhaps more similar than anticipated.