Different binding of NF-Y transcriptional factor to DQA1 promoter variants.
Polymorphism in the HLA-DQA1 promoter (QAP) sequences could influence the gene expression through a differential binding of transcriptional factors. Considering the main role played by the Y-box in the transcription, we focused on the QAP4 variants differing for a G vs A transition from the QAP Y-box consensus sequence. Electrophoretic Mobility Shift Assay using the two Y-box sequences was performed to determine whether this mutation could be reflected in an allele-specific binding of transcriptional factors. Indeed, the NF-Y specific band, recognised by supershift experiments, was clearly observed using the Y-box consensus probe but it was barely detectable with the QAP4 one. On the contrary, two other complexes were found to more strongly interact with QAP4 Y-box in comparison to the consensus sequence. The analysis of a selected panel of HLA homozygous lymphoblastoid cell lines by competitive RT-PCR and by Northern blotting revealed that the DQA1 *0401, *0501,*0601 alleles regulated by the QAP4 promoters were less expressed at the mRNA level than the DQA1* 0201 allele regulated by the QAP2.1 variant. In conclusion, these results show an evident reduction of NF-Y binding to the mutated QAP4 Y-box and a decreased mRNA accumulation of the DQA1 alleles regulated by these variants.
Indovina
P
P
Department of Experimental Medicine and Pathology, La Sapienza University of Rome, Italy.
Megiorni
F
F
Ferrante
P
P
Apollonio
I
I
Petronzelli
F
F
Mazzilli
M C
MC
D001483:Base Sequence; D015152:Blotting, Northern; D022762:CCAAT-Enhancer-Binding Proteins; D002461:Cell Line, Transformed; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005802:Genes, MHC Class II / Q000235:genetics; D014644:Genetic Variation; D006683:HLA-DQ Antigens / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012333:RNA, Messenger / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment; D014157:Transcription Factors / Q000378:metabolism*
D022762:CCAAT-Enhancer-Binding Proteins; D004268:DNA-Binding Proteins; D006683:HLA-DQ Antigens; D012333:RNA, Messenger; D014157:Transcription Factors