Transcriptional Deregulation of Homeobox Gene ZHX2 in Hodgkin Lymphoma

Abstract 2629

Hodgkin/Reed-Sternberg (HRS) cells represent the malignant fraction of infiltrated lymph nodes in Hodgkin lymphoma (HL). Although HRS cells display multiple chromosomal aberrations, few are recurrent and the targeted genes unknown. However, understanding the pathology of HL and developing rational therapies may well require identification and characterization of these genes and their deregulating mechanisms. Recently, we have identified a novel chromosomal rearrangement in HL cell line L-1236, t(4;8)(q27;q24), which targets ZHX2 at the recurrent breakpoint 8q24. ZHX2 encodes a transcription factor of the homeobox gene family and possesses tumour suppressor activity connected with a worse prognosis in HL-related multiple myeloma. Comparative expression analysis in HL cell lines and primary hematopoietic cells indicated aberrant downregulation in HL as well. In L-1236 the translocation breakpoint at 8q24 deletes the regulative far upstream region of ZHX2, indicating loss of activating elements. Therefore, we have looked for potential binding sites within this deleted region to analyze deregulation of this B-cell tumour suppressor gene. Accordingly, we identified two transcription factors, homeodomain protein MSX1 and bZIP protein XBP1, regulating ZHX2 expression. SiRNA-mediated knockdown and reporter gene analyses indicated that both factors activated transcription of ZHX2 directly via their corresponding binding sites. Furthermore, MSX1-cofactor histone H1C mediated repression of ZHX2 and showed enhanced expression levels in L-1236. As analyzed by fluorescence in situ hybridization (FISH) and genomic arrays, the gene loci of MSX1 at 4p16 and H1C at 6p21 were rearranged in several HL cell lines, correlating with their altered expression activity. As compared to primary hematopoietic cells, XBP1 expression was reduced in 6/7 HL cell lines while one cell line showed elevated levels, corresponding to a chromosomal rearrangement therein. Interestingly, the neighbouring pro-apoptotic genes EDEM1 and BHLHE40 were removed by del(3p26) in this cell line and activated by XBP1 in other HL cells. Taken together, our results demonstrate multiple mechanisms decreasing expression of tumour suppressor gene ZHX2 in HL cells: loss of enhancing binding sites, reduced expression of activators MSX1 and XBP1, and overexpression of MSX1-corepressor H1C. Moreover, chromosomal deregulations of genes involved in this regulative network highlight their role in development and malignancy of B-cells.