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. 2011 Mar 17;471(7338):377-81.
doi: 10.1038/nature09754. Epub 2011 Mar 2.

MHC class II transactivator CIITA is a recurrent gene fusion partner in lymphoid cancers

Christian Steidl  1 , Sohrab P Shah, Bruce W Woolcock, Lixin Rui, Masahiro Kawahara, Pedro Farinha, Nathalie A Johnson, Yongjun Zhao, Adele Telenius, Susana Ben Neriah, Andrew McPherson, Barbara Meissner, Ujunwa C Okoye, Arjan Diepstra, Anke van den Berg, Mark Sun, Gillian Leung, Steven J Jones, Joseph M Connors, David G Huntsman, Kerry J Savage, Lisa M Rimsza, Douglas E Horsman, Louis M Staudt, Ulrich Steidl, Marco A Marra, Randy D Gascoyne
Affiliations

MHC class II transactivator CIITA is a recurrent gene fusion partner in lymphoid cancers

Christian Steidl et al. Nature. .

Abstract

Chromosomal translocations are critically involved in the molecular pathogenesis of B-cell lymphomas, and highly recurrent and specific rearrangements have defined distinct molecular subtypes linked to unique clinicopathological features. In contrast, several well-characterized lymphoma entities still lack disease-defining translocation events. To identify novel fusion transcripts resulting from translocations, we investigated two Hodgkin lymphoma cell lines by whole-transcriptome paired-end sequencing (RNA-seq). Here we show a highly expressed gene fusion involving the major histocompatibility complex (MHC) class II transactivator CIITA (MHC2TA) in KM-H2 cells. In a subsequent evaluation of 263 B-cell lymphomas, we also demonstrate that genomic CIITA breaks are highly recurrent in primary mediastinal B-cell lymphoma (38%) and classical Hodgkin lymphoma (cHL) (15%). Furthermore, we find that CIITA is a promiscuous partner of various in-frame gene fusions, and we report that CIITA gene alterations impact survival in primary mediastinal B-cell lymphoma (PMBCL). As functional consequences of CIITA gene fusions, we identify downregulation of surface HLA class II expression and overexpression of ligands of the receptor molecule programmed cell death 1 (CD274/PDL1 and CD273/PDL2). These receptor-ligand interactions have been shown to impact anti-tumour immune responses in several cancers, whereas decreased MHC class II expression has been linked to reduced tumour cell immunogenicity. Thus, our findings suggest that recurrent rearrangements of CIITA may represent a novel genetic mechanism underlying tumour-microenvironment interactions across a spectrum of lymphoid cancers.

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Conflict of interest statement

The authors declare no competing financial interests.

Figures

Figure 1
Figure 1. CIITA–BX648577 gene fusion observed using paired-end massively parallel whole transcriptome sequencing
In the upper panel, 468 mate-pair sequences are shown aligning on either side of the breakpoint (pairing CIITA and BX648577). The genomic coordinates of the exon boundaries are given. In light blue, the transcribed intronic BX648577 sequence is shown as part of a transcript variant resulting from the fusion. In the lower panel, 191 split-reads are depicted lying on the breakpoint (in blue: merged reference sequence of CIITA and BX648577). The histogram on the right describes the absolute frequency of each sequence read spanning the breakpoint.
Figure 2
Figure 2. Molecular characterization of the gene fusion CIITA–BX648577 in Hodgkin lymphoma cell line KM-H2
a, Genomic location: exon structure and genomic breakpoints. b, Genomic fusion: rearranged genomic location with fusion of CIITA exons 1–5 with intron 1 of BX648577. c, Fusion transcript: the longest fusion transcript with transcribed intronic BX648577 sequence (*) is shown. Shorter splice variants exist. d, Reading frame at the breakpoint and putative translation: CIITA exons 1–4 and BX648577 exon 2 original reading frames are conserved. The shorter splice variants leading to premature translational stop at the breakpoint are not shown. e, BX648577 gene expression: gene expression array data (Affymetrix HG UA133 2.0 Plus probe set ID 243309_at) showing overexpression of BX648577 in KM-H2 compared with microdissected germinal centre B cells and other Hodgkin lymphoma cell lines. f, BX648577 exon-specific expression is biased towards exon 2 as part of the CIITA–BX648577 gene fusion. g, RNA interference with the gene fusion in KM-H2 cells increases surface HLA-DR expression compared with the non-silencing control. h, Forced expression of the CIITA–BX648577 fusion decreases surface HLA-DR expression on SUDHL4 cells compared with empty vector controls. Mean fluorescence intensities (ym) are indicated.
Figure 3
Figure 3. FISH on tissue microarrays showing recurrent CIITA break-apart in cHL and PMBCL
Representative images are shown. a, Design of the break-apart assay using bacterial artificial chromosome probes RP11-109M19-SpG (green signals) plus RP11-66H6-SpO (red signals). b, Combined immunofluorescence for CD30 (red staining) and FISH shows CIITA rearrangement in Hodgkin Reed–Sternberg cells that carry the CD30 antigen. Upper image: Hodgkin Reed–Sternberg cell-rich area with CIITA break-apart, lower images: individual Hodgkin Reed–Sternberg cells with break-apart. c, PMBCL case with CIITA break-apart in almost all cells represented in the section. Signal constellation indicates CIITA polyploidy and rearrangement of multiple alleles. d, Disease-specific survival of 57 patients with PMBCL treated with multi-agent chemotherapy (with or without radiation) according to CIITA rearrangement status. The presence of a CIITA rearrangement significantly correlated with shorter disease-specific survival (P = 0.044)
Figure 4
Figure 4. CD274/CD273 expression on PMBCL cells inhibits T-cell activation
a, mRNA overexpression of CD274 and CD273 in molecular subtypes of DLBCL including PMBCL. Normalized relative log2 ratios are shown (Affymetrix gene expression profiling). Probeset intensities of CD274 and CD273 correlate with each other (Pearson coefficient 0.541). b, Fold change of CD274 and CD273 mRNA expression in PMBCL compared with germinal centre B cells including cases with CIITA–CD274 (*), CIITA–CD273 (**) fusions and cell line U2940. c, Flow cytometric analysis of CD274/CD273 expression. Expression levels of CD274 or CD273 (blue histogram) and isotype controls (red histogram) are shown. d, Inhibition of T-cell activation by PMBCL cells. Dot plots and bar graph show that CD69 expression on Jurkat T cells (CD2 positive) is increasingly reduced by co-incubation with increasing numbers of U2940 cells. e, T-cell inhibition is PD1 dependent. CD274-Fc, CD273-Fc, PD1-Fc, CTLA4-Fc or control Ig were added to the co-cultures, respectively, in which Jurkat T cells were mixed with U2940 cells. Mean fluorescence intensities (MFI) are shown. Each bar is the mean of triplicate cultures, the error bars indicating the standard deviation (Student’s t-test, *P < 0.05, **P < 0.01, compared with control Ig). f, Forced expression of CIITA–CD274 and CIITA–CD273 fusion transcripts in U2932. The top panel shows the surface protein expression in transduced cells compared with empty vector control cells. The middle and bottom panels show significantly reduced CD69 expression on Jurkat T cells and IL-2 levels in the supernatant after admixture of CIITA–CD274 and −CD273 fusion-expressing U2932 cells. Each bar is the mean of triplicate cultures, the error bars indicating the standard deviation (Student’s t-test, *P < 0.05, **P < 0.01, compared with the empty vector control).
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