Abstract

Primary Sjögren's Syndrome (PSS) is a highly prevalent autoimmune disease, typically manifesting as lymphocytic infiltration of the exocrine glands leading to chronically impaired lacrimal and salivary secretion. Sjögren's Syndrome nuclear autoantigen 1 (SSNA1 or NA14) is a major specific target for autoantibodies in PSS but the precise function and clinical relevance of this protein are largely unknown. Orthologues of the gene are absent from many of the commonly used model organisms but are present in Chlamyodomonas reinhardtii (in which it has been termed DIP13) and most protozoa. We report the functional characterisation of the orthologue of SSNA1 in the kinetoplastid parasite, Trypanosoma brucei. Both TbDIP13 and human SSNA1 are small coiled-coil proteins which are predicted to be remote homologues of the actin-binding protein tropomyosin. We use comparative proteomic methods to identify potential interacting partners of TbDIP13. We also show evidence that TbDIP13 is able to self-assemble into fibril-like structures both in vitro and in vivo, a property which may contribute to its immunogenicity. Endogenous TbDIP13 partially co-localises with acetylated α-tubulin in the insect procyclic stage of the parasite. However, deletion of the DIP13 gene in cultured bloodstream and procyclic stages of T. brucei has little effect on parasite growth or morphology, indicating either a degree of functional redundancy or a function in an alternative stage of the parasite life cycle.

Highlights

  • Primary Sjogren’s Syndrome (PSS) is a highly prevalent autoimmune disease, affecting an estimated 0.5% of the population of the Western World

  • Identification of a T. brucei DIP13 orthologue As part of an ongoing study on N-myristoylated proteins in kinetoplastids, we identified a protein sequence encoded by the T. brucei brucei genome (Tb10.61.2720 from GeneDB Version 2.1), of which the C-terminus shared 33% identity with the Chlamydomonas reinhardtii flagellar protein DIP13 [11]

  • Overexpression of TbDIP13 in BSF parasites In order to investigate the subcellular localization of TbDIP13, C-terminal epitope-tagged forms of TbDIP (TbDIP13GFP and TbDIP13myc) were overexpressed using a tetracycline-inducible system in T. brucei bloodstream form (BSF) parasites (Figure 1A– D)

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Summary

Introduction

Primary Sjogren’s Syndrome (PSS) is a highly prevalent autoimmune disease, affecting an estimated 0.5% of the population of the Western World. Key features of this chronic disorder are lymphocytic infiltration of the exocrine glands leading to impaired lacrimal and salivary secretion, and the production of autoantibodies. The protein has been identified as a binding partner of the microtubule-severing protein spastin, a member of the AAA ATPase family, which is encoded by the SPG4 gene [8]. SSNA1 has been identified as a binding partner for the G-protein coupled receptor TPRA40, the expression of which is rapidly downregulated during hypoxia and reoxygenation in mammals [6]

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