PT - JOURNAL ARTICLE AU - Luis M. de Pablos AU - Tiago R. Ferreira AU - Adam A. Dowle AU - Sarah Forrester AU - Ewan Parry AU - Katherine Newling AU - Pegine B. Walrad TI - The mRNA-bound proteome of <em>Leishmania mexicana</em>: novel genetic insight into an ancient parasite AID - 10.1101/592402 DP - 2019 Jan 01 TA - bioRxiv PG - 592402 4099 - http://biorxiv.org/content/early/2019/03/28/592402.short 4100 - http://biorxiv.org/content/early/2019/03/28/592402.full AB - Leishmania parasite infections, termed the leishmaniases, cause significant global infectious disease burden. The lifecycle of the parasite embodies three main stages that require precise coordination of gene regulation to survive environmental shifts between sandfly and mammalian hosts. Constitutive transcription in kinetoplastid parasites means that gene regulation is overwhelmingly reliant on post-transcriptional mechanisms, yet strikingly few Leishmania trans-regulators are known. Utilizing optimised crosslinking and deep, quantified mass spectrometry, we present a comprehensive analysis of 1,400 mRNA binding proteins (mRBPs) and whole cell proteomes from the three main Leishmania lifecycle stages. Supporting the validity, while the crosslinked RBPome is magnitudes more enriched the protein identities of the crosslinked and non-crosslinked RBPomes were nearly identical. Moreover, multiple candidate RBPs were endogenously tagged and found to associate with discrete mRNA target pools in a stage-specific manner. Results indicate that in L.mexicana parasites, mRNA levels are not a strong predictor of the whole cell expression or RNA binding potential of encoded proteins. Evidence includes a low correlation between transcript and corresponding protein expression and stage-specific variation in protein expression versus RNA binding potential. Unsurprisingly, RNA binding protein enrichment correlates strongly with relative replication efficiency of the specific lifecycle stage. Our study is the first to quantitatively define and compare the mRBPome of multiple stages in kinetoplastid parasites. It provides novel, in-depth insight into the trans-regulatory mRNA:Protein (mRNP) complexes that drive Leishmania parasite lifecycle progression.