Global identification of conserved post-transcriptional regulatory programs in trypanosomatids

Abstract

While regulatory programs are extensively studied at the level of transcription, elements that are involved in regulation of post-transcriptional processes are largely unknown, and methods for systematic identification of these elements are in early stages. Here, using a novel computational framework, we have integrated sequence information with several functional genomics data sets to characterize conserved regulatory programs of trypanosomatids, a group of eukaryotes that almost entirely rely on post-transcriptional processes for regulation of mRNA abundance. This analysis revealed a complex network of linear and structural RNA elements that potentially govern mRNA abundance across different life stages and environmental conditions. Furthermore, we show that the conserved regulatory network that we have identified is responsive to chemical perturbation of several biological functions in trypanosomatids. We have further characterized one of the most abundant regulatory RNA elements that we discovered, an AU-rich element (ARE) that can be found in 3′ untranslated region of many trypanosomatid genes. Using bioinformatics approaches as well as in vitro and in vivo experiments, we have identified three ELAV-like homologs, including the developmentally critical protein TbRBP6, which regulate abundance of a large number of trypanosomatid ARE-containing transcripts. Together, these studies lay out a roadmap for characterization of mechanisms that modulate development and metabolic pathways in trypanosomatids.

Gene expression regulatory networks in Trypanosoma brucei: insights into the role of the mRNA‐binding proteome

“…The 25 canonical RBPs include PUF, RRM, CCCH and PSP1 domain‐containing proteins. Two canonical RBPs previously shown to negatively affect gene expression, RBP10 (Wurst et al ., ) and DRBD12 (Najafabadi et al ., ) are in this list. Not surprisingly, five subunits of the CAF1/NOT deadenylase complex reproducibly suppressed PGKB expression suggesting these subunits recruit the unique catalytic subunit CAF1 to the associated reporter mRNA (Farber et al ., ; Erben et al ., ).…”

The DRBD13 RNA binding protein is involved in the insect‐stage differentiation process of Trypanosoma brucei

“…Culturing and transfection of trypanosomes (Lister 427 strain expressing the tetracycline (tet) repressor from pHD1313 and 29–13) were performed as described in [20].…”

The emerging role of RNA-binding proteins in the life cycle ofTrypanosoma brucei

“…The metacyclic cells produced in vitro by inducible RBP6 expression were shown to express the VSG coat and importantly, the activated VSG genes possessed a metacyclic‐type VSG Pol I promoter and were found in monocistronic transcription units (Kolev et al ., ). More recently, it was suggested that RBP6 is a remote ELAV‐like T. brucei homologue that binds in vitro to an AU‐rich element (AUUUAUU) present in the 3′UTRs of mRNAs (Najafabadi et al ., ). Interestingly, expression of RBP6 in procyclic cells was found to downregulate mRNAs containing the identified sequence signature mRNAs (Najafabadi et al ., ), an opposite effect in comparison to available data for ELAV family proteins from other species.…”

“…More recently, it was suggested that RBP6 is a remote ELAV‐like T. brucei homologue that binds in vitro to an AU‐rich element (AUUUAUU) present in the 3′UTRs of mRNAs (Najafabadi et al ., ). Interestingly, expression of RBP6 in procyclic cells was found to downregulate mRNAs containing the identified sequence signature mRNAs (Najafabadi et al ., ), an opposite effect in comparison to available data for ELAV family proteins from other species. Identifying the precise in vivo RNA binding sites for RBP6 will undoubtedly clarify the repertoire of RBP6 targets and shed more light on its mode of action.…”