Biomedical subjects

L A Baglia

Publications and source records attributed to L A Baglia.

8 recordsLinked to original sources

PubMed1999

The E6 protein of human papillomavirus type 16 binds to and inhibits co-activation by CBP and p300.

The co-activators CBP and p300 are important for normal cell differentiation and cell cycle progression and are the targets for viral proteins that dysregulate these cellular processes. We show here that the E6 protein from the oncogenic human papillomavirus type 16 (HPV-16) binds to three regions (C/H1, C/H3 and the C-terminus) of both CBP and p300. The interaction of E6 with CBP/p300 was direct and independent of proteins known to bind the co-activators, such as p53. The E6 protein from low-risk HPV type 6 did not interact with C/H3 or the C-terminus but associated with the C/H1 domain at 50% of the level of HPV-16. HPV-16 E6 inhibited the intrinsic transcriptional activity of CBP/p300 and decreased the ability of p300 to activate p53- and NF-kappaB-responsive promoter elements. Interestingly, some mutations in HPV-16 E6 abrogated C/H3-E6 interactions, but did not alter the ability of E6 to associate with the C/H1 domain, suggesting that these modified proteins could be used to delineate the functional significance of the C/H1 and C/H3 domains of CBP/p300.

Animals↗

PubMed1998

Rb binds c-Jun and activates transcription.

The retinoblastoma protein (Rb) acts as a critical cell-cycle regulator and loss of Rb function is associated with a variety of human cancer types. Here we report that Rb binds to members of the AP-1 family of transcription factors, including c-Jun, and stimulates c-Jun transcriptional activity from an AP-1 consensus sequence. The interaction involves the leucine zipper region of c-Jun and the B pocket of Rb as well as a C-terminal domain. We also present evidence that the complexes are found in terminally differentiating keratinocytes and cells entering the G1 phase of the cell cycle after release from serum starvation. The human papillomavirus type 16 E7 protein, which binds to both c-Jun and Rb, inhibits the ability of Rb to activate c-Jun. The results provide evidence of a role for Rb as a transcriptional activator in early G1 and as a potential modulator of c-Jun expression during keratinocyte differentiation.

Animals↗

PubMed1997

The avian C/EBPgamma gene encodes a highly conserved leucine zipper transcription factor.

The C/EBP family of transcription factors regulates viral and cellular CCAAT/enhancer element-mediated transcription. We report the isolation and characterization of genomic and cDNA clones encoding avian CCAAT/enhancer-binding protein-gamma (C/EBP gamma). A partial cDNA clone for a C/EBP-related gene was previously identified by expression library screening for proteins binding the A1 CCAAT/enhancer motif from the avian leukosis virus long terminal repeat [W. Bowers and A. Ruddell (1992) J. Virol. 66, 6578-6586]. Additional cDNA and genomic clones were generated and sequenced to identify the complete protein coding sequence of this gene. Sequence analysis indicates that this gene encodes the avian homolog of C/EBP gamma. As with the murine C/EBP gamma homolog, the avian C/EBP gamma gene is comprised of two exons, with the open reading frame encoded in exon 2. The 150-aa C/EBP gamma protein is highly conserved, as the avian protein shows more than 80% identity with the murine and human homologs. The sequence of the initiation methionine (-3 caaAUGa +4) from the 150-aa open reading frame has a non-optimal Kozak initiation sequence. In vitro transcription and translation assay of this avian cDNA followed by radioimmunoprecipitation assay using a murine C/EBP gamma antiserum indicates that this non-optimal initiation codon is used to express a 22-kDa DNA-binding protein.

Amino Acid Sequence↗

PubMed1996

Regulation of avian leukosis virus long terminal repeat-enhanced transcription by C/EBP-Rel interactions.

The avian leukosis and sarcoma virus long terminal repeat (LTR) enhancers feature directly repeated CCAAT/enhancer element sequences which are also found in many viral and cellular gene enhancers. While most members of the CCAAT/enhancer element-binding protein (C/EBP) transcription factor family exhibit tissue-restricted expression, there may be ubiquitously expressed C/EBP-like factors that regulate widespread CCAAT/enhancer element-driven transcription. An avian C/EBP-related factor designated Al/EBP was previ- ously shown to bind CCAAT/enhancer elements within the avian leukosis virus (ALV) and Rous sarcoma virus (RSV) LTR enhancers in a pattern identical to that of a B-cell LTR-binding factor (W. J. Bowers and A. Ruddell, J. Virol. 66:6578-6586, 1992). An Al/EBP-specific antiserum recognizes a 40-kDa LTR CCAAT/enhancer element-binding protein purified from avian B lymphoma cells. A1/EBP is widely expressed at the mRNA and protein levels, suggesting that this protein could be important not only in regulating widespread expression of the AIN and RSV retroviruses but also in controlling the expression of other viral and cellular gene enhancers that possess CCAAT/enhancer motifs. We also found that an NF-KB/Rel-related protein is a component of the LTR CCAAT/enhancer element binding complex through its interaction with A1/EBP. At least one of the NF-kappaB family members, p65 (RelA), is capable of activating LTR CCAAT/enhancer element-driven transcription. These findings suggest a role for Rel-related factors in the regulation of AIN or RSV LTR-driven transcription via an interaction with Al/EBP.

Alpharetrovirus↗

PubMed1994

The VBP and a1/EBP leucine zipper factors bind overlapping subsets of avian retroviral long terminal repeat CCAAT/enhancer elements.

Two long terminal repeat (LTR) enhancer-binding proteins which may regulate high rates of avian leukosis virus (ALV) LTR-enhanced c-myc transcription during bursal lymphomagenesis have been identified (A. Ruddell, M. Linial, and M. Groudine, Mol. Cell. Biol. 9:5660-5668, 1989). The genes encoding the a1/EBP and a3/EBP binding factors were cloned by expression screening of a lambda gt11 cDNA library from chicken bursal lymphoma cells. The a1/EBP cDNA encodes a novel leucine zipper transcription factor (W. Bowers and A. Ruddell, J. Virol. 66:6578-6586, 1992). The partial a3/EBP cDNA clone encodes amino acids 84 to 313 of vitellogenin gene-binding protein (VBP), a leucine zipper factor that binds the avian vitellogenin II gene promoter (S. Iyer, D. Davis, and J. Burch, Mol. Cell. Biol. 11:4863-4875, 1991). Multiple VBP mRNAs are expressed in B cells in a pattern identical to that previously observed for VBP in other cell types. The LTR-binding activities of VBP, a1/EBP, and B-cell nuclear extract protein were compared and mapped by gel shift, DNase I footprinting, and methylation interference assays. The purified VBP and a1/EBP bacterial fusion proteins bind overlapping but distinct subsets of CCAAT/enhancer elements in the closely related ALV and Rous sarcoma virus (RSV) LTR enhancers. Protein binding to these CCAAT/enhancer elements accounts for most of the labile LTR enhancer-binding activity observed in B-cell nuclear extracts. VBP and a1/EBP could mediate the high rates of ALV and RSV LTR-enhanced transcription in bursal lymphoma cells and many other cell types.

Amino Acid Sequence↗

PubMed1992

Production of immunoreactive forms of growth hormone by the Burkitt tumor serum-free cell line sfRamos.

Tumor B-cell mitogenesis may be regulated by autocrine secretions that cross-react immunologically with neuroendocrine peptides. This hypothesis is supported by our recent report that immunoreactive human PRL (ir-hPRL) is produced by and required for the continuous growth of sfRamos, a Burkitt tumor serum-free cell line. Further support for the hypothesis is provided in this study. The data illustrate that antiserum immunoglobulin fraction G (IgG) to synthetic human GH (NIDDK anti-synth hGH IC-4), but not nonimmune serum IgG, completely inhibited sfRamos proliferation. In addition, anti-synth hGH, but not nonimmune serum, identified several polypeptides in sfRamos spent medium that were not in serum-free control medium, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) under reducing conditions and the Western immunoblot procedure. The ir-hGH polypeptides were in the 35- to more than 300-kilodalton (kDa) range, with a 48-kDa polypeptide as a major component. A polypeptide with electrophoretic properties identical to those of the hGH monomer (22 kDa) was not detected, even with prior incubation of spent medium with N-ethylmaleimide and dithiothreitol. Pituitary preparations of hPRL and hGH, each with the monomer form as the major component, were neither mitogenic for growth-arrested cells nor did they synergize with spent medium to enhance growth, either alone or in combination, whereas spent medium with immunoreactive polypeptides larger than the monomer form markedly stimulated proliferation of growth-arrested cells. These findings demonstrate that sfRamos mitogenesis is potentially regulated by autocrine secretions with cross-immunoreactivity to two anterior pituitary hormones.

Blotting, Western↗

PubMed1991

An Epstein-Barr virus-negative Burkitt lymphoma cell line (sfRamos) secretes a prolactin-like protein during continuous growth in serum-free medium.

Pituitary human PRL (hPRL) antiserum inhibits growth of B-lymphoblastoid cells in vitro, but the mechanism of inhibition is unclear. In this study the mechanism of inhibition of human B-cell growth by anti-hPRL was explored with an Epstein-Barr virus nuclear antigen (EBNA)-negative Burkitt lymphoma cell line (sfRamos) that proliferates continuously in serum-free medium with human transferrin as the only protein supplement. The data show that antiserum immunoglobulin fraction G (IgG) to pituitary hPRL, but not nonimmune serum IgG, completely inhibited the growth of sfRamos cells. In addition, anti-hPRL IgG identified a single band (29 kDa) in sfRamos spent medium, but not in fresh serum-free medium or in human transferrin, as demonstrated by sodium dodecyl sulfate-reducing polyacrylamide gel electrophoresis and Western immunoblot analysis. Polyacrylamide gel electrophoresis and Western analysis of a mixture containing sfRamos spent medium and excess pituitary hPRL established that the sfRamos 29-kDa PRL-like protein (PLP29) and pituitary hPRL (23 kDa) were electrophoretically distinct. Finally, sfRamos spent medium, but not fresh serum-free medium, was mitogenic for sfRamos and Nb2, a PRL-sensitive node rat lymphoma cell line. These findings demonstrate that PLP29 is biologically and immunologically like pituitary hPRL, but is electrophoretically different from this hormone. We suggest that PLP29 is secreted as an autocrine growth factor by sfRamos Burkitt lymphoma cells during continuous serum-free growth.

Burkitt Lymphoma↗

PubMed1988

Maintenance of growth transformation with Epstein-Barr virus is mediated by secretion of autocrine growth factors in two serum-free B-cell lines.

The characteristics of two tamarin (Saguinus oedipus) B-cell lines (sfBIT and sfBT) growth-transformed by Epstein-Barr virus (EBV) that proliferate continuously in serum-free medium are described. sfBIT was established by selecting cells for growth in RPMI 1640 supplemented with insulin, transferrin, and selenium (J. E. Shaw, R. G. Petit, and K. Leung, J. Virol. 61:4033-4037, 1987). sfBT, a subline of sfBIT cells reported here for the first time, required transferrin as the only protein supplement for continuous growth in RPMI 1640. Growth of sfBT cells was linear with human transferrin at 10(-2) to 10 micrograms/ml. Transferrin at 5 micrograms/ml yielded a culture density of 5 X 10(5) to 1 X 10(6) cells per ml, a cell doubling time of 2 to 3 days, and a culture viability greater than 95%. sfBIT and sfBT cells released transforming virus during continuous growth in serum-free culture medium without EBV-inducing agents. The spent medium of both serum-free lines supported cell growth at low culture density (1 x 10(4) to 5 X 10(4) cells per ml), but growth was arrested at low culture density with fresh serum-free medium. A procedure to measure growth-promoting activity (GPA) was established, and it revealed that the GPA of spent medium was greater than that of fresh medium for both serum-free cell lines. When fresh and spent media were dialyzed (molecular weight cutoff, 3,500) and subsequently concentrated by lyophilization, only the GPA of spent medium increased. We conclude that maintenance of growth transformation of tamarin cells latently infected with EBV is mediated by growth factors that are entirely autocrine in origin.

Animals↗