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Profile of Caprin-1 RNA-targets

Author(s):
Natacha Azussa Migita
Total Authors: 1
Document type: Master's Dissertation
Institution: Universidade Estadual de Campinas. Instituto de Biologia
Defense date:
Examining board members:
Cleslei Fernando Zanelli; Jörg Kobarg
Advisor: Katlin Brauer Massirer
Abstract

Caprin-1 (also known as RNG-105) is a cytoplasmic RNA-binding protein (RBP) expressed in mammalian tissues and is particularly abundant in neurons as part of RNA granules. This RBP binds mRNA targets in mRNPs and contains a conserved RG-rich domain, which is related to cellular granules and to reversible aggregation of proteins. Ectopic expression (EE) of Caprin-1 induces assembly of high-density cytoplasmic granules, called stress granules. Because activity and aggregation of RBPs are associated with many neurodegenerative and protein-misfolding diseases, we wanted to know the molecular mechanism by which Caprin-1 mediates stress granule induction and understand its function, focusing on the group of its RNA-targets. Here, we use human HEK293T cells to perform large-scale analysis including: RNA-immunoprecipitation and sequencing (RIP-seq) and RNA-sequencing (RNA-seq) upon Caprin-1 EE. RIP-seq results show 63% of rRNAs and 23% of protein-coding genes enriched in Caprin-1 RIP-seq in relation to control. Among the subgroup of RNA coding genes, most of the target candidates represent constituent of ribosome and components of translational initiation machinery. We have confirmed a handful of Caprin-1 transcripts-target involved in ribosome and translation initiation complex as 18S rRNA and NPM1. Interestingly, here we show that Caprin-1 protein auto-regulates its expression, but the molecular mechanisms have still to be elucidated. Through co-IP, we validated interactions of Caprin-1 with RPS3 and hnRNPC1/C2, ribosomal 40S subunit protein and a RBP that act to stabilize transcripts, respectively. By EE experiments, Caprin-1 is neither a universal repressor nor a universal activator, but some classes involved in translation machinery have been overepresented as repressed in the GO analysis. The combination of our results add to a link between Caprin-1 in stress granules and RNA processing. In this view, we propose that Caprin-1-RNA interactions might modulate stress granule and translation regulation and we expect to contribute to the understanding of its broader involved in neurodegenerative diseases. The next step is investigating the cellular and mechanistic understanding of the Caprin-1-dependent stress granules, since the physiological significance of most of the identified RNA targets has not yet been investigated. (AU)

FAPESP's process: 14/20174-6 - Implementation of CLIP-seq technique and evaluation of mRNAs-target by protein Caprin-1
Grantee:Natacha Azussa Migita
Support type: Scholarships in Brazil - Master