The TRIPLE PHD FINGERS proteins are required for SWI/SNF complex-mediated +1 nucleosome positioning and 5’ transcript length determination in Arabidopsis

Abstract

Eukaryotes have evolved multiple ATP-dependent chromatin remodelers to shape the nucleosome landscape. We recently uncovered an evolutionarily conserved SWItch/Sucrose Non-Fermentable (SWI/SNF) chromatin remodeler complex in plants reminiscent of the mammalian BAF subclass, which specifically incorporates the MINUSCULE (MINU) catalytic subunits and the TRIPLE PHD FINGERS (TPF) signature subunits. Here we report experimental evidence that establishes the functional relevance of TPF proteins for the complex activity. Our results show that depletion of TPF triggers similar pleiotropic phenotypes and molecular defects to those found in minu mutants. Moreover, we report the genomic location of MINU2 and TPF proteins as representative members of the plant BAF-like complex and their impact on nucleosome positioning and transcription. These analyses unravel the binding of the complex to thousands of genes where it modulates the position of the +1 nucleosome. These targets tend to produce 5′-shifted transcripts in the tpf and minu mutants pointing to the participation of the complex in alternative transcriptional start site usage. Interestingly, there is a remarkable correlation between +1 nucleosome shift and 5′ transcript length change suggesting their functional connection. In summary, this study unravels the function of a plant SWI/SNF complex involved in +1 nucleosome positioning and 5′ transcript length determination.

Organization, genomic targeting and assembly of three distinct SWI/SNF chromatin remodeling complexes inArabidopsis

“…Finally, consistent with the recent report (Diego-Martin et al, 2022), the MINUcontaining complexes do not contain the GIF2 subunit (SS18 paralogs) found in both BRM-and SYD-containing complexes, incorporate selective paralogs (that is, SWI3A/B but not SWI3C and SWI3D) and include a number of complex-specific subunits, including BSH, BRD5, SHH2, TPF1/2, OPF1/2, PSA1, and PSA2.…”

“…In contrast, the SYD-containing complexes selectively contain paralog SWI3D but not SWI3A/B/C, and include a complexspecific subunit, encoded by SYD sub-SWI/SNF Interacting Protein 1/2/3 (SSIP1, SSIP2, or SSIP3). Finally, consistent with the recent report (Diego-Martin et al, 2022), the MINU-containing complexes do not contain the GIF2 subunit (SS18 paralogs) found in both BRM-and SYD-containing complexes, incorporate selective paralogs (that is, SWI3A/B but not SWI3C and SWI3D) and include a number of complexspecific subunits, including BSH, BRD5, SHH2, TPF1/2, OPF1/2, PSA1, and PSA2.…”

“…For example, for SAS complexes, the three SSIPs (SSIP1, SSIP2, SSIP3) are separately incorporated into the SAS complexes (Figure 1B). Similarly, OPF1, the MAS-specific subunit, did not exist in the OPF2-containing MAS complexes (Diego-Martin et al, 2022). Finally, BASspecific subunit BRIP1 cannot co-immunoprecipitated with BRIP2, and BRD1 cannot co-immunoprecipitated with BRD2 and BRD13 (Figure 1B).…”

“…In addition, we recently biochemically isolated novel subunits of the BRM-containing SWI/SNF complexes in Arabidopsis, including two BRMinteracting proteins (BRIP1 and BRIP2) and three bromodomain-containing proteins, BRD1, BRD12, and BRD13 (Yu et al, 2020;Yu et al, 2021). More recently, MINUcontaining SWI/SNF complexes were also reported (Diego-Martin et al, 2022). Plant SWI/SNF subunits serve critical roles in cell differentiation, development, and response to various environmental signals, and their mutations result in severe defects in leaf development, root stem cell maintenance, flower patterning and timing, embryo development, and vegetative to adult phase transition (Sarnowski et al, 2005;Ho and Crabtree, 2010;Li et al, 2015;Wu et al, 2015;Yang et al, 2015;Zhao et al, 2015;Sarnowska et al, 2016;Xu et al, 2016).…”

The prefoldin-like protein AtURI exhibits characteristics of instrinsically disordered proteins

“…To prepare the pAtURI:AtURI1-3xFLAG construct, a genomic fragment starting 1312 nucleotides upstream of the AtURI ATG until the codon before the stop was amplified by PCR and a pENTR207-gAtURI was generated by Gateway. gAtURI was transferred into pEarleyGate302 containing a 3xFLAG tag [32] by Gateway. This construct and the pEarleyGate104-AtURI construct to overexpress YFP-AtURI were introduced into Col-0 WT Arabidopsis plants by Agrobacterium-mediated transformation.…”