Molecular and Cellular Pathobiology Epigenetic Regulator Smchd1 Functions as a Tumor Suppressor Huei San Leong 1,2 , Kelan Chen 1,2 , Yifang Hu 1 , Stanley Lee 1,2 , Jason Corbin 1 , Miha Pakusch 1 , James M. Murphy 1,2 , Ian J. Majewski 5 , Gordon K. Smyth 1,3 , Warren S. Alexander 1,2 , Douglas J. Hilton 1,2 , and Marnie E. Blewitt 1,2,4 Abstract SMCHD1 is an epigenetic modifier of gene expression that is critical to maintain X chromosome inactivation. Here, we show in mouse that genetic inactivation of Smchd1 accelerates tumorigenesis in male mice. Loss of Smchd1 in transformed mouse embryonic fibroblasts increased tumor growth upon transplantation into immunodeficient nude mice. In addition, loss of Smchd1 in Em-Myc transgenic mice that undergo lymphomagenesis reduced disease latency by 50% relative to control animals. In premalignant Em-Myc transgenic mice deficient in Smchd1, there was an increase in the number of pre-B cells in the periphery, likely accounting for the accelerated disease in these animals. Global gene expression profiling suggested that Smchd1 normally represses genes activated by MLL chimeric fusion proteins in leukemia, implying that Smchd1 loss may work through the same pathways as overexpressed MLL fusion proteins do in leukemia and lymphoma. Notably, we found that SMCHD1 is underexpressed in many types of human hematopoietic malignancy. Together, our observations collectively highlight a hitherto uncharacterized role for SMCHD1 as a candidate tumor suppressor gene in hematopoietic cancers. Cancer Res; 73(5); 1591–9. Ó2012 AACR. Introduction Smchd1 was originally identified as a regulator of the epi- genetic silencing of transgenes and metastable epialleles in mice (1). An ENU-induced point mutation, termed MommeD1 (MD1), results in nonsense-mediated mRNA decay, effectively producing a null allele of Smchd1. Smchd1 MD1/MD1 female embryos die in mid-gestation by embryonic day 10.5 (E10.5), whereas homozygous males are unaffected at this time. These observations were confirmed using a genetrap allele of Smchd1 (Smchd1 gt ), which also behaves as a null allele (2). Female- specific embryonic lethality is due to the critical role that Smchd1 plays in X chromosome inactivation, where Smchd1 is required for DNA methylation and silencing of genes on the inactive X chromosome and appears to be involved in the maintenance of X chromosome inactivation (2). Notably, only half of Smchd1 MD1/MD1 males survive to weaning (1), suggest- ing that Smchd1 has additional roles, consistent with its function in silencing autosomal transgenes and metastable epialleles. Smchd1 encodes a 2007 amino acid protein with an N- terminal ATP-binding domain and a conserved C-terminal SMC hinge domain. SMC hinge domains are otherwise found in the 6 canonical SMC family members, where a central hinge domain mediates DNA binding and heterodimeriza- tion between SMC proteins to form 3 distinct complexes involved in chromosome condensation, cohesion, and DNA repair (3). Many epigenetic regulators are aberrantly expressed or somatically mutated in diverse human cancers (4), so we were interested to test whether Smchd1 played a role in malignancy. Because of embryonic lethality of Smchd1-null females, we restricted our analyses to male mice. We show that loss of Smchd1 enhances growth in a fibroblast trans- formation model and accelerates development of Em-Myc B-cell lymphoma, showing that Smchd1 acts as a tumor suppressor. Materials and Methods Mouse strains Mice with mutations in Smchd1 (MommeD1 and genetrap alleles) were previously described (2). The Smchd1 MD1 mice were maintained on the FVB/N background, and Smchd1 gt mice were backcrossed with C57BL/6 mice for more than 10 generations. The Em-Myc transgenic mice were described previously (5) and are congenic with C57BL/6. All genotyping was as previously described (2, 6). All mice were housed under specific pathogen-free (SPF) conditions, under the approval of Authors' Affiliations: 1 The Walter and Eliza Hall Institute of Medical Research, Parkville; Departments of 2 Medical Biology, 3 Mathematics and Statistics, and 4 Genetics, University of Melbourne, Carlton, Victoria, Aus- tralia; and 5 The Netherlands Cancer Institute, Amsterdam, The Netherlands. Note: Supplementary data for this article are available at Cancer Research Online (http://cancerres.aacrjournals.org/). Corresponding Author: Marnie E. Blewitt, The Walter and Eliza Hall Institute of Medical Research, 1G Royal Pde, Parkville 3052 VIC, Australia. Phone: 61393452545; Fax: 61393470852; E-mail: blewitt@wehi.edu.au doi: 10.1158/0008-5472.CAN-12-3019 Ó2012 American Association for Cancer Research. Cancer Research www.aacrjournals.org 1591 on June 13, 2020. © 2013 American Association for Cancer Research. cancerres.aacrjournals.org Downloaded from Published OnlineFirst December 26, 2012; DOI: 10.1158/0008-5472.CAN-12-3019