Introduction of exogenous class II trans-activator in MHC class II-deficient ABI fibroblasts results in incomplete rescue of MHC class II antigen expression.
Previously, we have shown that fibroblasts established from type III bare lymphocyte syndrome patient ABI are characterized by the absence of MHC class II gene expression and a strongly reduced amount of MHC class I transcripts. Complementation analysis has suggested that the gene defective in these ABI fibroblasts is different from that encoding the class II trans-activator (CIITA), which has been attributed an essential role in both constitutive and inducible expression of MHC class II genes. In the present study it is shown by reverse transcriptase-PCR analysis that the amount of CIITA transcripts in ABI fibroblasts is greatly reduced compared with that in fibroblasts derived from a healthy individual. Transient cotransfection of a construct in which CIITA is under the control of a constitutive promoter with an HLA-DRA promoter-luciferase reporter plasmid resulted in enhanced luciferase expression in ABI fibroblasts. Furthermore, ABI fibroblasts stably transfected with CIITA re-express functional HLA-DR Ags, but do not express HLA-DQ and DP Ags at the cell surface. Comparison of these data with those obtained for normal fibroblasts and fibroblasts defective for CIITA indicate that the gene defect and the resulting lack of MHC class II expression in ABI fibroblasts can only partly be corrected by the introduction of CIITA. Furthermore, DNase I hypersensitivity analysis of ABI fibroblasts has revealed a closed chromatin structure in the promoter region of the MHC class II DRA gene. However, CIITA transfection resulted in an open DNA configuration, which suggests a role for CIITA in provoking changes in the chromatin structure of the DRA gene.
Peijnenburg
A
A
Department of Immunohematology and Blood Bank, University Hospital Leiden, The Netherlands.
Gobin
S J
SJ
van Eggermond
M C
MC
Godthelp
B C
BC
van Graafeiland
N
N
van den Elsen
P J
PJ
D002461:Cell Line, Transformed; D005347:Fibroblasts / Q000276:immunology; D005434:Flow Cytometry; D005786:Gene Expression Regulation; D018014:Gene Transfer Techniques; D000949:Histocompatibility Antigens Class II / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D006801:Humans; D009687:Nuclear Proteins; D015534:Trans-Activators / Q000235:genetics*
D000949:Histocompatibility Antigens Class II; C083728:MHC class II transactivator protein; D009687:Nuclear Proteins; D015534:Trans-Activators