Biomedical subjects

K G Becker

Publications and source records attributed to K G Becker.

39 records · Page 3Linked to original sources

PubMed1993

Retinoic acid induction of major histocompatibility complex class I genes in NTera-2 embryonal carcinoma cells involves induction of NF-kappa B (p50-p65) and retinoic acid receptor beta-retinoid X receptor beta heterodimers.

Retinoic acid (RA) treatment of human embryonal carcinoma (EC) NTera-2 (NT2) cells induces expression of major histocompatibility complex (MHC) class I and beta-2 microglobulin surface molecules. We found that this induction was accompanied by increased levels of MHC class I mRNA, which was attributable to the activation of the two conserved upstream enhancers, region I (NF-kappa B like) and region II. This activation coincided with the induction of nuclear factor binding activities specific for the two enhancers. Region I binding activity was not present in undifferentiated NT2 cells, but binding of an NF-kappa B heterodimer, p50-p65, was induced following RA treatment. The p50-p65 heterodimer was produced as a result of de novo induction of p50 and p65 mRNAs. Region II binding activity was present in undifferentiated cells at low levels but was greatly augmented by RA treatment because of activation of a nuclear hormone receptor heterodimer composed of the retinoid X receptor (RXR beta) and the RA receptor (RAR beta). The RXR beta-RAR beta heterodimer also bound RA responsive elements present in other genes which are likely to be involved in RA triggering of EC cell differentiation. Furthermore, transfection of p50 and p65 into undifferentiated NT2 cells synergistically activated region I-dependent MHC class I reporter activity. A similar increase in MHC class I reporter activity was demonstrated by cotransfection of RXR beta and RAR beta. These data show that following RA treatment, heterodimers of two transcription factor families are induced to bind to the MHC enhancers, which at least partly accounts for RA induction of MHC class I expression in NT2 EC cells.

Animals↗

PubMed1993

A regulatory element in the beta 2-microglobulin promoter identified by in vivo footprinting.

Expression of the beta 2-microglobulin (beta 2-m) and major histocompatibility complex (MHC) class I genes is coordinately regulated. By ligation-mediated polymerase chain reaction, we have analyzed in vivo factor binding to the promoter region of the murine beta 2-m gene. In adult spleen, in which beta 2-m is expressed, strong protection was found in three elements. Two of these elements, the beta 2-m NF-kappa B binding site and the interferon consensus sequence, are homologous to the regulatory elements of the MHC class I genes and were also found to be protected in spleen. A third protected element, PAM, identified in this work, is unique to the beta 2-m gene. None of the elements showed protection in brain tissue, in which neither the beta 2-m nor the MHC class I gene is expressed. In vivo footprinting was also performed with F9 embryonal carcinoma cells, in which expression of the beta 2-m and MHC class I genes is induced at a low level only upon stimulation with retinoic acid (RA). No in vivo protection was detected before and after RA treatment of F9 cells, indicating that RA induction of beta 2-m (and MHC class I) expression occurs without detectable in vivo factor occupancy, whereas EL4 T lymphocytes expressing beta 2-m at a high level exhibited strong protection similar to that in spleen. Despite the lack of in vivo occupancy, the nuclear factors specific for each of the three elements were present in brain tissue and F9 cells as well as in spleen tissue and EL4 cells. We show that PAM, an element identified by its in vivo protection, binds nuclear factors ranging from 40 to 50 kDa in size and is capable of enhancing transcription of a reporter in F9 and other cells. Taken together, these results indicate that in vivo factor occupancy for the beta 2-m and MHC class I promoters is coordinated and occurs through a mechanism other than mere expression of relevant factors.

Animals↗

PubMed1992

Cloning of a negative transcription factor that binds to the upstream conserved region of Moloney murine leukemia virus.

The long terminal repeat of Moloney murine leukemia virus (MuLV) contains the upstream conserved region (UCR). The UCR core sequence, CGCCATTTT, binds a ubiquitous nuclear factor and mediates negative regulation of MuLV promoter activity. We have isolated murine cDNA clones encoding a protein, referred to as UCRBP, that binds specifically to the UCR core sequence. Gel mobility shift assays demonstrate that the UCRBP fusion protein expressed in bacteria binds the UCR core with specificity identical to that of the UCR-binding factor in the nucleus of murine and human cells. Analysis of full-length UCRBP cDNA reveals that it has a putative zinc finger domain composed of four C2H2 zinc fingers of the GLI subgroup and an N-terminal region containing alternating charges, including a stretch of 12 histidine residues. The 2.4-kb UCRBP message is expressed in all cell lines examined (teratocarcinoma, B- and T-cell, macrophage, fibroblast, and myocyte), consistent with the ubiquitous expression of the UCR-binding factor. Transient transfection of an expressible UCRBP cDNA into fibroblasts results in down-regulation of MuLV promoter activity, in agreement with previous functional analysis of the UCR. Recently three groups have independently isolated human and mouse UCRBP. These studies show that UCRBP binds to various target motifs that are distinct from the UCR motif: the adeno-associated virus P5 promoter and elements in the immunoglobulin light- and heavy-chain genes, as well as elements in ribosomal protein genes. These results indicate that UCRBP has unusually diverse DNA-binding specificity and as such is likely to regulate expression of many different genes.

Amino Acid Sequence↗