Identification of a Nuclear Export Sequence in the MHC CIITA

Abstract

Initiation of an immune response through expression of MHC class II and related genes is under the control of the CIITA. Normally found in both the cytoplasm and nucleus, CIITA is tightly controlled by a variety of posttranslational modifications as well as interactions with other nuclear and cytoplasmic factors, whereas disruption of this dual subcellular localization impairs CIITA functioning and expression of target genes. Although CIITA has well-defined domains necessary for its nuclear import, the region responsible for the translocation of CIITA from the nucleus has not been characterized. In this study, we identify a leucine-rich motif at residues 717–724 that bears strong homology to known nuclear export sequence (NES) domains. Mutation of this region renders CIITA insensitive to treatment with leptomycin B, an inhibitor of nuclear export, whereas fusion of this domain to a heterologous GFP is sufficient to induce its export to the cytoplasm or cause its retention in the nucleus following leptomycin B treatment. Point mutations of specific leucine residues within the NES disrupt the normal subcellular distribution of the full-length CIITA, impair its ability to interact with the nuclear export factor CRM1, and enhance CIITA-induced gene expression from an MHC class II gene promoter. IFN-γ stimulation of class II genes is further enhanced by inhibiting the nuclear export of endogenous CIITA. Collectively, these data demonstrate the first identification of a specific NES within CIITA and place it among the other protein domains that contribute to the posttranslational regulation of CIITA activity.

HLA Class II Antigen Processing and Presentation Pathway Components Demonstrated by Transcriptome and Protein Analyses of Islet β-Cells From Donors With Type 1 Diabetes

“…Immunohistological analysis verified that the small complement of b-cells expressing Class II also stain for CIITA, which was mainly located in the cytoplasm rather than in the nucleus. Nuclear CIITA has been visualized using green fluorescent protein or FLAGtagged CIITA (35,36), or by using sufficient nuclear protein for Western blot detection. However, we were unable to detect CIITA protein in the nucleus of endocrine or immune cells in pancreas tissue sections, in human tonsillar tissue sections ( Supplementary Fig.…”

Detection and prognostic value of recurrent exportin 1 mutations in tumor and cell-free circulating DNA of patients with classical Hodgkin lymphoma

“…Several proteins known to play a major role in cHL oncogenesis, including STAT1/STAT6, FOXO1 or CIITA are also identified as cargo proteins. [34][35][36] Whether the XPO1 mutations may interfere with the nucleus/cytoplasm shuttle of these proteins remains to be determined.…”

David and Goliath: chemical perturbation of eukaryotes by bacteria

“…The specificity of leptomycin B has been used to validate the CRM1-dependent export of many NES- containing proteins including actin [ 181 ], cytokines [ 140 ], tyrosine kinases [ 169 ], cyclin-CDK [ 75 , 191 ], MDM2/p53 [ 64 ], inhibitors of NF-κB transcription [ 155 ] and MHC class II complexes [ 36 ]. Inhibition by this drug results in the accumulation of these key regulatory proteins which eventually leads to cell death.…”