A Vestigial Pathway for Sex Differences in Immune Regulation

RESEARCH HIGHLIGHT

A vestigial pathway for sex differences in immune regulation

Yun Liang1, J Michelle Kahlenberg2 and Johann E Gudjonsson1

Cellular & Molecular Immunology (2017) 14, 578–580; doi:10.1038/cmi.2017.28; published online 22 May 2017

utoimmune diseases have a strik- until they noticed the small ‘growth’ on primary androgenic hormone, testoster- Aingly higher prevalence in women the female skin. one, can stimulate development of char- compared to men, with approximately From the perspective of evolution, acteristics used in sexual selection while 80% of individuals affected with auto- sexual dimorphisms are often explained increasing risks of infection owing to its immune diseases being female. The nat- by sexual selection, the selection arising immunosuppressive effects.5 Indeed, tes- ure of this sex-bias has remained unclear, from differential mating success. In the tosterone has been shown to reduce but sex hormones have previously been example of the anglerfish, it is thought natural killer-cell activity; expression of implicated as the leading underlying that in the sparsely populated ocean, the Toll-like receptors (TLR); and synthesis of cause. In this research highlight, we chances for a male to find his mate are so tumornecrosisfactor(TNF),inducible discuss a recently identified transcription low that once this happens, he attaches nitric oxide synthase and nitrogen factor, vestigial like family member 3 to her and never leaves. In this way he oxide by macrophages.6–8 Autoimmune (VGLL3), as a non-hormonally influ- can provide her with sperm his entire regulator (AIRE), a key factor in central enced regulator of the heightened inflam- life, and in return he gets all his nutrients tolerance that promotes self-antigen matory responses observed in women, from her. Without the need to hunt for expression in medullary thymic epithelial and discuss its role as a potential driver food, he degenerates all his internal cells, has been shown to be upregulated by of autoimmunity. organs except the testes. androgens and downregulated by Sexual dimorphism, in which different Sex differences in humans, although estrogen.9,10 However, intriguingly, in characteristics are exhibited by the two arguably subtler than those in anglerfish, humans, mutations in AIRE, which sexes of the same species, has long fasci- are important in medicine. Drugs can cause autoimmune polyendocrinopathy- nated scientists. One remarkable example require sex-specific dosing, and their candidiasis-ectodermal dystrophy, are is the anglerfish. While females measure effects can differ between men and characterized by combinations of auto- approximately 10 cm in length, in some women.1 Many diseases are more pre- immune diseases such as Addison’sdis- subordersthemalescanbe10times valent in one sex, with autoimmune ease, vitiligo, type I diabetes and alopecia, smaller, with the record being 6.2 mm. diseases being one of the most striking which are diseases that do not generally The males are so tiny, living attached to examples. Systemic lupus erythematosus show gender bias.11 females, that scientists wondered why all (SLE) has a female-to-male ratio of 9:1. To understand the mechanism of sex the anglerfish they captured were females The female-to-male ratio of scleroderma differences in susceptibility to immune and Sjögren’s syndrome can be as high as diseases, we have investigated sexual 20:1, and that of Grave’s disease can dimorphisms in immune processes on a reach 7:1.2 Overall, it is estimated that molecular level.12 Using high-resolution 1Department of Dermatology, University of Michi- 78% of people affected with autoim- transcriptome analyses, we found 661 gan, Ann Arbor, MI 48109, USA and 2Division of mune diseases are women.2 By contrast, genes that are expressed differentially by Rheumatology, Department of Internal Medicine, infectious diseases are more prevalent in the two sexes in human skin. The University of Michigan, Ann Arbor, MI 48109, men.3 female-biased genes (that is, genes USA Correspondence: Dr JE Gudjonsson, Department Sexual selection has similarly operated expressed more highly in females than of Dermatology, University of Michigan, 6427 during human evolution, as supported by in males) were significantly enriched for Med Sci I 1300 E. Catherine St., Ann Arbor, MI sex differences in attributes such as phy- processes known to be dysregulated in 48109, USA. sical size that are related to advantages in autoimmune diseases, including comple- E-mail: [email protected] − Received: 30 March 2017; Accepted: 3 April male male competition and female ment activation and phagocytosis regula- 2017 choice.4 It has been proposed that the tion. By contrast, the male-biased genes A vestigial pathway for sex differences YLianget al

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Is the role of VGLL3 in autoimmune regulation restricted to the skin? In Sjögren’s syndrome, an autoimmune condition of the salivary and lacrimal glands that disproportionally affects more women than men at a 20:1 ratio,19 the expression of VGLL3 and its targets was increased. Many sexually Figure 1 VGLL3-regulated gene network promotes autoimmunity. The female-biased dimorphic autoimmune diseases have expression and nuclear localization of VGLL3 leads to elevated expression of pro- in common a prominent activation of autoimmune genes, which may help explain the increased susceptibility of females to both type I and type II interferon (IFN) autoimmune diseases. signaling. Notably, VGLL3 disruption by RNAi impaired the response of cultured were enriched for transcription regula- gene vg (‘vestigial’), which encodes a salivary gland cells to IFN-α and IFN-γ. tion and development. Importantly, the cofactor of Scalloped,ahomologofthe The regulation of interferon responses by female-biased genes were significantly transcriptional enhancer TEF-1.16 Nota- VGLL3 is conserved in monocytes as associated with common disease loci of bly, VGLL3 in salmon exhibits sex- well, as VGLL3 knockdown reduced the SLE and systemic sclerosis, two female- dependent dominance, promoting earlier induction of bona fide type I interferon dominant autoimmune diseases. The and later maturation in males and response genes. This suggests that one female-increased genes included BAFF/ females, respectively.17 This suggests that mechanism by which VGLL3 promotes TNFSF13B, the target for the first biolo- the role of VGLL3 in regulating sexual development of autoimmune diseases is gic therapy approved for SLE, and dimorphism is evolutionarily conserved. through heightened interferon responses. ITGAM, whose variants are associated In primary human keratinocytes, Since the discovery of sex differences with SLE susceptibility from genome- VGLL3 knockdown reduced the expres- in susceptibility to immune-associated wide association studies.13,14 sion of 7 out of 10 keratinocyte-expressed diseases, many studies have suggested Further analysis of the data demon- genes that are associated with female- the differential regulation of immune strated a genome-wide co-expression biased diseases. On a genome-wide level, genes such as TLRs, CCLs, CXCLsand network that extended beyond the 661 all of the top ten biological pathways ILs, in the two sexes.3 These studies have sex-biased genes. This network involved enriched in VGLL3 targets are related to further suggested that hormonal and multiple inflammatory pathways, indi- immune processes. In addition, ‘autoim- genetic mediators on the sex chromo- cating the presence of extensive mune diseases’ was among the top disease somes modulate these differences.3 We genome-wide differences in immune categories enriched, involving 47% of have extended these findings and estab- regulation between the two sexes. VGLL3-regulated genes. lished a significant association between Interestingly, we demonstrated that In subacute cutaneous lupus erythe- sex-biased immune gene expression in sex hormones do not affect the expres- matosus (SCLE), a female-biased, the human skin and disease loci for sion of the female-biased genes that are lupus-specific skin eruption, the autoimmunity. In addition, we have involved in immune processes. Consis- disease-upregulated genes showed a demonstrated that the expression of tently, the expression of these genes did strong correlation with VGLL3- these genes, the majority of which are not decrease with age, despite a decrease regulated genes. Furthermore, VGLL3 located on autosomes, are not affected by in estrogen levels after menopause. These knockdown in SCLE keratinocytes sig- sex hormones and biological age. The findings raise the possibility that sex-bias nificantly reversed the abnormality in identification of VGLL3 as their upstream in immune regulation is independent of gene expression. Notably, active disease regulator provides a new, sex hormone- sex hormones and is further corrobo- was associated with increased VGLL3 independent perspective to the molecular rated by various clinical studies showing protein expression and nuclear localiza- basis of sex disparities in immune gene lack of association between the onset or tion, consistent with increased activation regulation (Figure 1). activity of autoimmune diseases with sex of this pathway in inflamed tissue. Simi- Decades ago, it was proposed that hormone levels.15 larly, transcriptional upregulation by men were more susceptible to infection By screening transcription factors that VGLL3 was associated with active states because their larger body size provided demonstrate significant female-bias in of morphea and systemic sclerosis, two more ‘bites’ to bacteria. Over the years, human skin, we identified VGLL3 as female-biased autoimmune diseases of we have made substantial progress in the only transcription factor that was the skin.18 Importantly, VGLL3 was understanding the mechanisms under- required for the expression of a large mostly cytoplasmic (transcriptionally lying sex biases in immune responses, set of female-biased immune genes, inactive) in healthy male skin, but in but we still have many more questions including BAFF/TNFSF13B, ITGAM males with SCLE, it was noted to be ahead of us to answer. For example, what and C3.HumanVGLL3 was originally nuclear and likely participatory in is the mechanism of gene regulation by identified as a homolog to the Drosophila disease. VGLL3? How is VGLL3 itself regulated

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in the two sexes? What is the role of 4 Geary DC. Sex differences in social behavior autoimmune diseases. Nat Immunol 2017; – VGLL3 in innate and adaptive immune and cognition: utility of sexual selection for 18:152 160. hypothesis generation. Horm Behav 2006; 13 Vincent FB, Morand EF, Schneider P, responses? By understanding sex as a 49:273–275. Mackay F. The BAFF/APRIL system in SLE biological variable in immune responses 5 Folstad I, Karter AJ. Parasites, bright males, pathogenesis. Nat Rev Rheumatol 2014; and the pathogenesis of immune dis- and the immunocompetence handicap. Am 10:365–373. – 14 International Consortium for Systemic Lupus eases, we may one day find a way to Nat 1992; 139:603 622. 6 Rettew JA, Huet-Hudson YM, Marriott I. Erythematosus Genetics (SLEGEN), Harley JB, successfully combat or even prevent Testosterone reduces macrophage expression Alarcón-Riquelme ME, Criswell LA, Jacob CO, these diseases. in the mouse of Toll-like receptor 4, a trigger Kimberly RP et al. Genome-wide asso- for inflammation and innate immunity. Biol ciation scan in women with systemic lupus fi Reprod 2008; 78:432–437. erythematosus identi es susceptibility CONFLICT OF INTEREST variants in ITGAM, PXK, KIAA1542 fl 7 Hou J, Zheng WF. Effect of sex hormones on The authors declare no con ict of interest. NK and ADCC activity of mice. Int J Immu- and other loci. Nat Genet 2008; 40: – nopharmacol 1988; 10:15–22. 204 210. ACKNOWLEDGEMENTS 8 D'Agostino P, Milano S, Barbera C, Di Bella G, 15 Holroyd CR, Edwards CJ. The effects of hormone replacement therapy on autoim- The work is supported by the US National La Rosa M, Ferlazzo V et al. Sex hormones modulate inflammatory mediators produced mune disease: rheumatoid arthritis and sys- Institutes of Health (K08-AR060802 and R01- temic lupus erythematosus. Climacteric by macrophages. Ann NY Acad Sci 1999; 2009; 12:378–386. AR069071to JEG; and R03-AR066337 and 876: 426–429. 16 Maeda T, Chapman DL, Stewart AF. Mam- K08-AR063668 to JMK), an A Alfred 9 Zhu ML, Bakhru P, Conley B, Nelson JS, malian vestigial-like 2, a cofactor of TEF-1 Taubman Medical Research Institute Kenneth Free M, Martin A et al. Sex bias in CNS and MEF2 transcription factors that pro- and Frances Eisenberg Emerging Scholar autoimmune disease mediated by androgen motes skeletal muscle differentiation. JBiol Award (JEG), the Doris Duke Charitable control of autoimmune regulator. Nat Com- Chem 2002; 277: 48889–48898. Foundation (2013106 to JEG) and a Pfizer mun 2016; 7: 11350. 17 Barson NJ, Aykanat T, Hindar K, Baranski M, 10 Dragin N, Bismuth J, Cizeron-Clairac G, Aspire Award (JEG). Bolstad GH, Fiske P et al. Sex-dependent Biferi MG, Berthault C, Serraf A et al. dominance at a single locus maintains Estrogen-mediated downregulation of AIRE variation in age at maturity in salmon. influences sexual dimorphism in autoim- Nature 2015; 528: 405-+. mune diseases. J Clin Invest 2016; 126: 18 Leitenberger JJ, Cayce RL, Haley RW, – 1 Clayton JA, Collins FS. Policy: NIH to bal- 1525 1537. Adams-Huet B, Bergstresser PR, Jacobe ance sex in cell and animal studies. Nature 11 Pitkanen J, Peterson P. Autoimmune regu- HT. Distinct autoimmune syndromes in mor- 2014; 509:282–283. lator: from loss of function to autoimmunity. phea: a review of 245 adult and 2 Whitacre CC. Sex differences in autoimmune Genes Immun 2003; 4:12–21. pediatric cases. Arch Dermatol 2009; 145: disease. Nat Immunol 2001; 2:777–780. 12 Liang Y, Tsoi LC, Xing X, Beamer MA, 545–550. 3 Klein SL, Flanagan KL. Sex differences in Swindell WR, Sarkar MK et al. Agene 19 Patel R, Shahane A. The epidemiology of immune responses. Nat Rev Immunol 2016; network regulated by the transcription factor Sjogren's syndrome. Clin Epidemiol 2014; 16:626–638. VGLL3 as a promoter of sex-biased 6:247–255.

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