Mip6 binds directly to the Mex67 UBA domain to maintain low levels of Msn2/4 stress dependent mRNAs

Abstract del trabajo presentado en 12ª Reunión de la Red Española de Levaduras. El Escorial, Madrid.11-13 de diciembre de 2019Pág. 44 del libro de abstracts que se adjunta. RNA-binding proteins (RBPs) participate in all steps of gene expression, underscoring their potential as regulators of RNA homeostasis. We structurally and functionally characterize Mip6, a four-RNA recognition motif (RRM)-containing RBP, as a functional and physical interactor of the export factor Mex67. Mip6-RRM4 directly interacts with the ubiquitin-associated (UBA) domain of Mex67 through a loop containing tryptophan 442. Mip6 shuttles between the nucleus and the cytoplasm in a Mex67-dependent manner and concentrates in cytoplasmic foci under stress. Photoactivatable ribonucleoside-enhanced crosslinking and immunoprecipitation experiments show preferential binding of Mip6 to mRNAs regulated by the stress-response Msn2/4 transcription factors. Consistent with this binding, MIP6 deletion affects their export and expression levels. Additionally, Mip6 interacts physically and/or functionally with proteins with a role in mRNA metabolism and transcription such as Rrp6, Xrn1, Sgf73, and Rpb1. These results reveal a novel role for Mip6 in the homeostasis of Msn2/4-dependent transcripts through its direct interaction with the Mex67 UBA domain

Mip6 binds directly to the Mex67 UBA domain to maintain low levels of Msn2/4 stress-dependent mRNAs

21 páginas, 8 figuras. PAR-CLIP datasets are available at GEO as GSE126236: https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE126236RNA-binding proteins (RBPs) participate in all steps of gene expression, underscoring their potential as regulators of RNA homeostasis. We structurally and functionally characterize Mip6, a four-RNA recognition motif (RRM)-containing RBP, as a functional and physical interactor of the export factor Mex67. Mip6-RRM4 directly interacts with the ubiquitin-associated (UBA) domain of Mex67 through a loop containing tryptophan 442. Mip6 shuttles between the nucleus and the cytoplasm in a Mex67-dependent manner and concentrates in cytoplasmic foci under stress. Photoactivatable ribonucleoside-enhanced crosslinking and immunoprecipitation experiments show preferential binding of Mip6 to mRNAs regulated by the stress-response Msn2/4 transcription factors. Consistent with this binding, MIP6 deletion affects their export and expression levels. Additionally, Mip6 interacts physically and/or functionally with proteins with a role in mRNA metabolism and transcription such as Rrp6, Xrn1, Sgf73, and Rpb1. These results reveal a novel role for Mip6 in the homeostasis of Msn2/4-dependent transcripts through its direct interaction with the Mex67 UBA domain.MM-E, CN-C and AT-C were supported by the GVA (Val I+D: ACIF/2015/025) PROMETEO (PROM/2016/093) and GVA (Val I+D: ACIF/2019/212). NM was partially supported by the European Union Erasmus Mundus programme. NMR experiments were performed in the "Manuel Rico" NMR laboratory (LMR) (CSIC). The authors would like to thank Diamond Light Source for beamtime (proposal mx-10121), and the staff of beamline I04 for assistance with crystal testing and data collection. This study was supported by funds to SR-N from the Spanish MINECO, MICIIN (BFU2014-57636, BFU2015-71978, PGC2018- 099872-B-I00) and the Generalitat Valenciana (PROM/2012/061, ACOMP2014/061 and PROMETEO 2016/093), and to JB from the Spanish MINECO (SAF2015-67077-R, SAF2017-89901-R) and JMP-C from the Spanish MINECO (CTQ2018-84371) and Comunidad de Madrid (B2017/BMD-3770).Peer reviewe