NLRC5 exclusively transactivates MHC class I and related genes through a distinctive SXY module

MHC class II (MHCII) genes are transactivated by the NOD-like receptor (NLR) family member CIITA, which is recruited to SXY enhancers of MHCII promoters via a DNA-binding “enhanceosome” complex. NLRC5, another NLR protein, was recently found to control transcription of MHC class I (MHCI) genes. However, detailed understanding of NLRC5’s target gene specificity and mechanism of action remained lacking. We performed ChIP-sequencing experiments to gain comprehensive information on NLRC5-regulated genes. In addition to classical MHCI genes, we exclusively identified novel targets encoding non-classical MHCI molecules having important functions in immunity and tolerance. ChIP-sequencing performed with Rfx5−/− cells, which lack the pivotal enhanceosome factor RFX5, demonstrated its strict requirement for NLRC5 recruitment. Accordingly, Rfx5-knockout mice phenocopy Nlrc5 deficiency with respect to defective MHCI expression. Analysis of B cell lines lacking RFX5, RFXAP, or RFXANK further corroborated the importance of the enhanceosome for MHCI expression. Although recruited by common DNA-binding factors, CIITA and NLRC5 exhibit non-redundant functions, shown here using double-deficient Nlrc5−/−CIIta−/− mice. These paradoxical findings were resolved by using a “de novo” motif-discovery approach showing that the SXY consensus sequence occupied by NLRC5 in vivo diverges significantly from that occupied by CIITA. These sequence differences were sufficient to determine preferential occupation and transactivation by NLRC5 or CIITA, respectively, and the S box was found to be the essential feature conferring NLRC5 specificity. These results broaden our knowledge on the transcriptional activities of NLRC5 and CIITA, revealing their dependence on shared enhanceosome factors but their recruitment to distinct enhancer motifs in vivo. Furthermore, we demonstrated selectivity of NLRC5 for genes encoding MHCI or related proteins, rendering it an attractive target for therapeutic intervention. NLRC5 and CIITA thus emerge as paradigms for a novel class of transcriptional regulators dedicated for transactivating extremely few, phylogenetically related genes.
Author Summary Major histocompatibility complex class I (MHCI) molecules are central to immunity and immunological disorders, and constitute a major obstacle in organ transplantation. It is therefore vital to gain insight into the regulation of their expression. NLRC5 was recently found to regulate MHCI gene transcription. However, we lack a thorough understanding of its target gene specificity and mechanism of action. Our work addresses these questions, delineating the unique consensus sequence required for NLRC5 recruitment and pinpointing conserved features conferring its specificity. Furthermore, through genome-wide analyses, we confirm that NLRC5 regulates classical MHCI genes and identify novel target genes, all encoding non-classical MHCI molecules exerting an array of functions in immunity and tolerance. We thereby demonstrate that NLRC5 exclusively transactivates genes of the MHCI pathway, rendering it an attractive target for future therapeutic intervention. The most striking feature of NLRC5 is its restricted and highly focused transcriptional activity, which has been described so far only for one related factor, CIITA. NLRC5 and CIITA therefore emerge as prototypes for a novel kind of extremely specific transcriptional regulator.