Kinetoplastid SL RNA biogenesis

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DESCRIPTION (provided by the applicant): Our research is aimed at understanding gene expression in the family Trypanosomatidae, which includes the parasitic protozoon's responsible for leishmaniasis, African Sleeping Sickness, and Chagas Disease. The focus is on the genesis and function of the spliced leader (SL) RNA, a small RNA that contributes the 5'-end sequence to every nuclear messenger RNA via a trans-splicing reaction. Trans-splicing, which is necessary for the conversion of polycistronic pre-messenger RNA (a likely consequence of the unusual genome organization found in kinetoplastids) into monocistronic mRNA, is not found in the human host or insect vector and thus represents a possible therapeutic target. This proposal outlines three complementary sets of experiments that detail the maturation pathway of the SL RNA common to Leishmania tarentolae and Trypanosoma brucei. 1) To refine the map of cis elements (SL RNA sequences and structures) necessary for accurate and efficient maturation and trans-splicing. Data obtained will allow the future identification of trans factors (protein and RNA) that interact with essential regions of RNA. 2) To isolate and examine the role of an XPO1-dependent nuclear export complex in SL RNA trafficking. 3) Experimental examination of the roles of a 3'-exonuclease and two ribose 2'-O- methyltransferases in the maturation of the 3' and 5' ends of the SL RNA, and bioinformatic identification and modeling of these proteins' interaction with the SL RNA. These three goals address discrete steps in the maturation of the SL RNA that determine its subsequent function. Specific inhibition of the trans-splicing pathway is lethal to the cell and thus a prime target for anti-parasite intervention.