Differential expression patterns of c-REL protein in classic and nodular lymphocyte predominant Hodgkin lymphoma.

Classic Hodgkin lymphoma (cHL) is characterized by numerical gains of the short arm of chromosome 2. The high frequency of 2p overrepresentation including REL, particularly in the nodular sclerosis subtype suggests that constitutive activation of nuclear factor kappaB/REL is a hallmark of Reed-Sternberg (RS) cells. The aim of this study was to investigate c-Rel protein expression patterns in cHL and nodular lymphocyte predominant Hodgkin lymphoma (NLPHL) cases by immunohistochemical analysis. A total of 79 cases of HL were analyzed, which included 59 cases of cHL (49 nodular sclerosis; 8 mixed cellularity; 2 lymphocyte-rich) and 20 cases of NLPHL. Positive staining was defined in this study as a reaction seen in the nuclei or nuclei and cytoplasm of RS or lymphocytic and histiocytic (L&H) cells in cHL and NLPHL cases, respectively. The percent positivity of c-REL staining of RS cells in cHL was seen in 51 of 59 cases (86.4%). No significant difference in c-REL expression was seen between nodular sclerosis (42 of 49, 85.7%) and mixed cellularity subtypes (7 of 8 cases, 87.5%; P = 1). In comparison, positive c-REL protein expression in L&H cells was seen in 5 of 20 NLPHL cases (25.0%). Therefore, significantly higher positivity of RS cells in cHL was seen compared with positivity of L&H cells in NLPHL; 86.4% vs. 25.0%; P = 0). Expression of Epstein-Barr virus latent membrane protein was seen in 6 of 30 cases (19.0%; 25 cHL, 5 NLPHL) and EBER1 in 5 of 27 cases (18.5%; 24 cHL, 3 NLPHL). The presence of Epstein-Barr virus did not correlate with c-REL protein expression (P = 1). Our results demonstrate that there is differential c-REL protein expression in cHL in comparison with NLPHL and suggest that c-REL may play a role in the pathogenesis of classic Hodgkin lymphoma.

Role of USF1 phosphorylation on cardiac alpha-myosin heavy chain promoter activity.

Contractile activity of the cardiac myocyte is required for maintaining cell mass and phenotype, including expression of the cardiac-specific alpha-myosin heavy chain (alpha-MHC) gene. An E-box hemodynamic response element (HME) located at position -47 within the alpha-MHC promoter is both necessary and sufficient to confer contractile responsiveness to the gene and has been shown to bind upstream stimulatory factor-1 (USF1). When studied in spontaneously contracting cardiac myocytes, there is enhanced binding of USF1 to the HME compared with quiescent cells, which correlates with a threefold increase in alpha-MHC promoter activity. A molecular mechanism by which contractile function modulates alpha-MHC transcriptional activity may involve signaling via phosphorylation of USF1. The present studies showed that purified rat USF1 was phosphorylated in vitro by protein kinase C (PKC) and cAMP-dependent protein kinase (PKA) but not casein kinase II. Phosphorylated USF1 by either PKC or PKA had increased DNA binding activity to the HME. PKC-mediated phosphorylation also leads to the formation of USF1 multimers as assessed by gel shift assay. Analysis of in vivo phosphorylated nuclear proteins from cultured ventricular myocytes showed that USF1 was phosphorylated, and resolution by two-dimensional gel electrophoresis identified at least two distinct phosphorylated USF1 molecules. These results suggest that endogenous kinases can covalently modify USF1 and provide a potential molecular mechanism by which the contractile stimulus mediates changes in myocyte gene transcription.

TSP50, a possible protease in human testes, is activated in breast cancer epithelial cells.

Initial studies have identified TSP50 as a human testes-specific gene that is demethylated in breast cancer. In this study, we will present new data related to the TSP50 gene. We have found that the TSP50 gene product shares a similar enzymatic structure with many serine proteases. However, the most critical catalytic site, serine, has been replaced by threonine. Western analysis revealed that in human testes, the TSP50 antibody detected two closely positioned protein bands whose estimated molecular masses were 37 kDa, whereas in a large portion of breast cancer tissues, but not normal control tissues, only one band was present. Immunohistochemistry assays found TSP50 proteins located in the spermatocytes of human testes, whereas in situ hybridization and immunohistochemistry confirmed that gene activation in breast tumors took place in malignant mammary epithelial cells. These results suggested that the normal function of the TSP50 gene was involved in spermatogenesis, whereas the up-regulation of TSP50 in many breast cancer patients not only indicated that it might be a novel biomarker for this disease but also encouraged us to further explore the possibility of whether it was an oncogene.