The Conserved Autoimmune-Disease Risk Gene TMEM39A Regulates Lysosome Dynamics
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The conserved autoimmune-disease risk gene TMEM39A regulates lysosome dynamics Shuo Luoa, Xin Wanga, Meirong Baia, Wei Jianga, Zhe Zhanga, Yifan Chenb, and Dengke K. Maa,c,1 aCardiovascular Research Institute, University of California, San Francisco, CA 94158; bDepartment of Molecular and Cell Biology, University of California, Berkeley, CA 94720; and cDepartment of Physiology, University of California, San Francisco, CA 94158 Edited by Victor R. Ambros, University of Massachusetts Medical School, Worcester, MA, and approved December 15, 2020 (received for review June 9, 2020) TMEM39A encodes an evolutionarily conserved transmembrane Appendix, Fig. S1B). In addition, tmem-39 mutant animals also protein and carries single-nucleotide polymorphisms associated have markedly higher expression of ER stress reporter hsp-4p::gfp with increased risk of major human autoimmune diseases, includ- in the hypoderm, supporting an important role of TMEM-39 in ing multiple sclerosis. The exact cellular function of TMEM39A re- normal protein homeostasis (Fig. 1D). To investigate if disruption mains not well understood. Here, we report that TMEM-39, the of TMEM-39 impairs the nervous system, the primary target of sole Caenorhabditis elegans (C. elegans) ortholog of TMEM39A, MS, we examined neuronal morphology of the mutant animals regulates lysosome distribution and accumulation. Elimination of using transgenic strains that express cholinergic (unc-17p::gfp), tmem-39 leads to lysosome tubularization and reduced lysosome GABAergic (unc-47p::gfp), or somatosensory PVD interneuron mobility, as well as accumulation of the lysosome-associated mem- (F49H12.4::gfp) reporters. About 10 to 20% of the wild-type brane protein LMP-1. In mammalian cells, loss of TMEM39A leads 8animals showed mild morphological abnormalities (e.g., short to redistribution of lysosomes from the perinuclear region to cell SI Appendix periphery. Mechanistically, TMEM39A interacts with the dynein filopodia-like structures) when they aged (Table 1 and , intermediate light chain DYNC1I2 to maintain proper lysosome Fig. S1G). Strikingly, disruption of tmem-39 ledtoseveremor- distribution. Deficiency of tmem-39 or the DYNC1I2 homolog in phological defects of the nervous system in 50 to 90% of the mutant C. elegans impairs mTOR signaling and activates the downstream animals examined, including long ectopic branches, incorrect neu- TFEB-like transcription factor HLH-30. We propose evolutionarily rite projections, and beading of neuronal processes (Fig. 1E and conserved roles of TMEM39 family proteins in regulating lysosome Table 1). We also found that the dendritic structures of the PVD distribution and lysosome-associated signaling, dysfunction of neurons were missing in L2 and L3 larva, despite the presence of CELL BIOLOGY which in humans may underlie aspects of autoimmune diseases. the PVD soma, suggesting developmental defects of the neuronal processes (SI Appendix,Fig.S1H). Together, our observations in- TMEM39A | lysosome | dynein | autoimmunity | autoimmune disorders dicate that TMEM-39 is important for maintaining normal ER homeostasis and neuronal morphology in the nervous system. utoimmune diseases affect millions of people worldwide, To determine the subcellular localization of TMEM-39 pro- Awhile exact causes of the diseases remain not well under- tein, we generated transgenic strains that express either a rescuing stood, despite decades of research (1–3). Recently, genome-wide tmem-39 genomic transgene with mCherry tagged to the N ter- association studies have identified single-nucleotide polymor- minus or N-mCherry–tagged tmem-39 complementary DNA phisms (SNPs) in TMEM39A that are associated with enhanced (cDNA) under an hsp-16.48 heat-shock promoter. The tmem-39 risks of multiple sclerosis (MS), systemic lupus erythematosus (SLE), and autoimmune thyroid disease (4–8), indicating an important link Significance between TMEM39A and autoimmune disease. TMEM39A is also overexpressed in glioma cell lines and glioma patient cells (9), im- Autoimmune diseases are characterized by overactivation of plicating its possible role in glioma pathogenesis. TMEM39A encodes an organism’s immune system and subsequent tissue damage a predicted multitransmembrane protein, homologs of which are and pathogenesis. Although genome-wide association studies found in most multicellular organisms (Fig. 1 A and B and SI Ap- have identified genetic variations, such as single-nucleotide pendix,Fig.S1A). A recent study reported that TMEM39A localizes polymorphisms (SNPs), that are linked to autoimmunity, the to the endoplasmic reticulum (ER) and regulates autophagy through exact function of the genes involved and molecular mecha- controlling trafficking of the PtdIns(4)P phosphatase SAC1 (10). To nisms underlying their contribution to autoimmunity remain ’ investigate TMEM39A s physiological roles and whether its cellular not well understood. Here, we show that an evolutionarily functions might be evolutionarily conserved, we generated Caeno- conserved autoimmunity risk gene, TMEM39A, regulates lyso- rhabditis elegans (C. elegans) strains and HEK 293T human cell lines some dynamics through interacting and functioning with mo- that lack respective TMEM39 homologs in each system. Phenotypic lecular motor dynein intermediate light chain. As lysosomes and protein-interactor analyses revealed conserved roles of TMEM39 are implicated in various aspects of immune-system dysfunc- in regulating lysosome dynamics and signaling, providing a plausible tion, including abnormal cytokine secretion and antigen pre- link of human TMEM39A to pathogenesis of autoimmune diseases. sentation, our study provides an important mechanistic insight Results suggesting how TMEM39A might contribute to autoimmunity like multiple sclerosis and lupus. We identified the uncharacterized C. elegans gene D1007.5 (named as tmem-39 hereafter) in a previous genome-wide RNA interference Author contributions: S.L. and D.K.M. designed research; S.L., X.W., M.B., W.J., Z.Z., Y.C., and (RNAi) screen for cellular-stress regulators (11). tmem-39 en- D.K.M. performed research; S.L. and D.K.M. analyzed data; and S.L. and D.K.M. wrote the paper. codes the only homolog of mammalian TMEM39A in the C. The authors declare no competing interest. elegans genome. Using CRISPR-Cas9, we either removed the This article is a PNAS Direct Submission. entire coding region of tmem-39 or inserted stop codons in the Published under the PNAS license. first exon of the gene (Fig. 1C). Mutant animals carrying tmem- 1To whom correspondence may be addressed. Email: [email protected]. 39 null alleles showed multiple developmental abnormalities, This article contains supporting information online at https://www.pnas.org/lookup/suppl/ including slow growth, reduced body length, and a tendency to doi:10.1073/pnas.2011379118/-/DCSupplemental. burst, indicating defects in cuticle development (Fig. 1D and SI Published February 2, 2021. PNAS 2021 Vol. 118 No. 6 e2011379118 https://doi.org/10.1073/pnas.2011379118 | 1of8 Downloaded by guest on October 1, 2021 A NH2 TMEM39A, human M Q V P R Q L G W T W Y C S G S L T G V P L G P S Q G R S T N R G R L C L N Y A K N P F R S S G G R Y H H A P N T N Y Y D M I L N L F R Q R S E K W N H L G G V S N I L H T W R T S S S A I Q D T C S K C L S L I L Y I Y L W L V N V E H L Y S A P F H A V L L F L L M G A G C L I Y L V P L V A I W C L L P F V Q F I F P L L F L T F L A M W V V L I A Y T M L G P I N S C Y T L I V L L Y F V Y A G N Y I F F A V L G Y F E I V P L R R Y A F V M L F T F E R A S V S L I L L G V F L W I V L L T T L K L P R L V A L P C S Q N L R H L S L V M C N F L L L Q G S Y H L L P R D C I D I E H F V S L Q E K P V A I P R Y D K Y D L P T M V G S H E W H V N F R I S E S V Q G L E L T S V C I C K Q Q I L R Y V Y L K S H V N L A G A A R Y H V A L K P C E L R D G K P W V H P A F F L L K A N E T K L E I P N K H L L L A S G S G V R G P A H Y A S S L Q P L R A N L H E N E Q R M P A D F R Y N L D V A K S F T S I H A H S N P C A S H A S COOH L T E S D L N A P L K A G Y T W L R D S Y A A to T in the risk allele rs1132200 L E F N D S S I Y V B Human Mouse Zebrafish Drosophila C. elegans C E dma268(S22ochre; R25opal) cholinergic GABAergic PVD D dma258(del) Bright field hsp-4p::gfp wild type wild type tmem-39(dm258) tmem-39(dma258) tmem-39(dma268) tmem-39(dma268) Fig.