Diversification of the Histone Acetyltransferase GCN5 through Alternative Splicing in Brachypodium distachyon

Abstract

The epigenetic modulatory SAGA complex is involved in various developmental and stress responsive pathways in plants. Alternative transcripts of the SAGA complex's enzymatic subunit GCN5 have been identified in Brachypodium distachyon. These splice variants differ based on the presence and integrity of their conserved domain sequences: the histone acetyltransferase domain, responsible for catalytic activity, and the bromodomain, involved in acetyl-lysine binding and genomic loci targeting. GCN5 is the wild-type transcript, while alternative splice sites result in the following transcriptional variants: L-GCN5, which is missing the bromodomain and S-GCN5, which lacks the bromodomain as well as certain motifs of the histone acetyltransferase domain. Absolute mRNA quantification revealed that, across eight B. distachyon accessions, GCN5 was the dominant transcript isoform, accounting for up to 90% of the entire transcript pool, followed by L-GCN5 and S-GCN5. A cycloheximide treatment further revealed that the S-GCN5 splice variant was degraded through the nonsense-mediated decay pathway. All alternative BdGCN5 transcripts displayed similar transcript profiles, being induced during early exposure to heat and displaying higher levels of accumulation in the crown, compared to aerial tissues. All predicted protein isoforms localize to the nucleus, which lends weight to their purported epigenetic functions. S-GCN5 was incapable of forming an in vivo protein interaction with ADA2, the transcriptional adaptor that links the histone acetyltransferase subunit to the SAGA complex, while both GCN5 and L-GCN5 interacted with ADA2, which suggests that a complete histone acetyltransferase domain is required for BdGCN5-BdADA2 interaction in vivo. Thus, there has been a diversification in BdGCN5 through alternative splicing that has resulted in differences in conserved domain composition, transcript fate and in vivo protein interaction partners. Furthermore, our results suggest that B. distachyon may harbor compositionally distinct SAGA-like complexes that differ based on their histone acetyltransferase subunit.

Genome-wide Analysis of Bromodomain Gene Family in Arabidopsis and Rice

“…While human Brd-homologs are important in several molecular mechanisms (Sanchez and Zhou, 2009; Sanchez et al, 2014; Filippakopoulos et al, 2012; Taniguchi, 2016; Cochran et al, 2019; Morgado-Pascual et al, 2019; Uppal et al, 2019; Boyson et al, 2021), only few plant Brd-homologs have been analyzed, including BRD-ET proteins of A. thaliana (GTE1/IMB1, Duque and Chua, 2003; GTE6, Chua et al 2005; GTE4, Airoldi et al, 2010; GTE9; GTE 11, Misra et al 2017), B. distachyon GCN5 (Martel, et al, 2017), and N. benthamiana NbDBCP (Sukarta et al 2020). The plant Brd-homologs with multi-domain architecture, are likely to be involved in diverse cellular mechanisms.…”

“…In both plants, the duplication and AS-events enhanced the heterogeneity of the Brd-homologs. AS-events were also shown to responsible for the differential fate and interaction of two GCN5 isoforms with similar localization (Martel et al, 2017). Therefore, understanding the significance of AS-events of At/OsBrds-homologs is important, as in certain cases AS-isoforms (e.g., OsBrd1.2, lacks BRD) are more abundant than the CS-isoforms, in both tissue- and stress-specific environment (Supplementary Figure 8).…”

“…Plants also harbour multiple Brd-gene members, however, unlike animals their functional significance is relatively less explored. Studies on few Brd-homologs from Arabidopsis and other plants have shown their involvement in seed germination (Duque and Chua 2003), leaf development (Chua et al 2005), mitotic cell cycle (Airoldi et al 2010), sugar and ABA responses (Misra et al 2017), development (Martel et al 2017; Rao et al 2014; Kalantidis et al 2007), and pathogen perception (Sukarta et al 2020). The Brd-gene family members show variable numbers in different organisms (Rao et al 2014).…”

Updated Mechanisms of GCN5—The Monkey King of the Plant Kingdom in Plant Development and Resistance to Abiotic Stresses

“…Additionally, alternative splicing in GCN5 transcripts have been reported in humans as well as Brachypodium distachyon (i.e., GCN5 , L-GCN5 , and S-GCN5 splice variants) [ 92 ], in which GCN5 and L-GCN5 but not S-GCN5 could interact with ADA2, indicating that these isoforms function in the diversification of the SAGA complex association, and shed light on the differences in the protein organization of plant GCN5-containing complexes [ 92 ]. Thus, plants may hold compositionally distinct SAGA-like complexes based on the different histone acetyltransferase subunits.…”

Blocking the Bromodomains Function Contributes to Disturbances in Alga Chara vulgaris Spermatids Differentiation

“…Plants studies have mainly focused on Arabidopsis , in which 29 BRD proteins have been identified [ 10 ] and functions of only three genes, from among 12, have been characterized [ 11 ]. Brachypodium [ 12 ], tomato and tobacco species [ 13 ] as well as soybean [ 10 ] have also been analyzed. Although researches related to plant bromodomains are ongoing, there is still no evidence to support the Florence and Faller [ 14 ] hypothesis explaining the presence of one bromodomain in plant proteins instead of two, as is the case in animals.…”