Chromosomal organization and evolutionary conservation of the human chromosome 19q linkage group containing three DNA repair genes

A series of human-rodent somatic cell hybrids were investigated by Southern blot analysis for the presence or absence of twenty-six molecular markers and three isozyme loci from human chromosome 19. Based on the co-retention of these markers in the various independent hybrid clones containing portions of human chromosome 19 and on pulsed field mapping, chromosome 19 is divided into twenty ordered regions. The most likely marker order for the chromosome is: (LDLR, C3)-(cen-MANNB)-D19S7-PEPD-D19S9-GPI-TGF$ \beta$-(CYP2A, NCA, CGM2, BCKAD)-PSG1a-(D19S8, XRCC1)-(D19S19, ATP1A3)-(D19S37, APOC2)-CKMM-ERCC2-ERCC1-(D19S62, D19S51)-D19S6-D19S50-D19S22-(CGB, FTL)-qter.^ The region of 19q between the proximal marker D19S7 and the distal gene coding for the beta subunit of chorionic gonadotropin (CGB) is about 37 Mb in size and covers about 37 cM genetic distance. The ration of genetic to physical distance on 19q is therefore very close to the genomic average OF 1 cM/Mb. Estimates of physical distances for intervals between chromosome 19 markers were calculated using a mapping function which estimates distances based on the number of breaks in hybrid clone panels. The consensus genetic distances between individual markers (established at HBM10) were compared to these estimates of physical distances. The close agreement between the two estimates suggested that spontaneously broken hybrids are as appropriate for this type of study as radiation hybrids.^ All three DNA repair genes located on chromosome 19 were found to have homologues on Chinese hamster chromosome 9, which is hemizygous in CHO cells, providing an explanation for the apparent ease with which mutations at these loci were identified in CHO cells. Homologues of CKMM and TGF$\beta$ (from human chromosome 19q) and a mini-satellite DNA specific to the distal region of human chromosome 19q were also mapped to Chinese hamster 9. Markers from 19p did not map to this hamster chromosome. Thus the q-arm of chromosome 19, at least between the genes PEPD and ERCC1, appears to be a linkage group which is conserved intact between humans and Chinese hamsters. ^

Cytochrome P450 4F isoforms in different species: Identification, gene regulation and putative roles in inflammation

The cytochrome P450 4F subfamily comprises a group of enzymes that metabolize derivatives of arachidonic acid such as prostaglandins, lipoxins leukotrienes and hydroxyeicosatetraenoic acids, which are important mediators involved in the inflammatory response. Therefore, we speculate that CYP4Fs might be able to modulate the extent of the inflammation by controlling of the tissue levels of these inflammatory mediators, especially, leukotriene B4. One way to provide support for this hypothesis is to test whether the expression of CYP4Fs changes under inflammatory conditions, since these changes are required to adjust the levels of inflammatory mediators. ^ A lipopolysacchride (LPS) induced rat inflammation model was used to analyze the expressions of rat CYP4F4 and CYP4F5 in liver and kidney. LPS administration did not change the constitutive expression level of CYP4F4 and CYP4F5. In liver, the expressions of CYP4F4 and CYP4F5 decreased to 50–60% of the untreated level. The same effect of LPS on CYP4F4 and CYP4F5 expression can be mimicked in hepatocyte primary cultures treated with LPS, indicating a direct of effect of LPS on hepatocytes. LPS treatment also decreased the activity of liver microsomes towards chlorpromazine, however, antibody inhibition study revealed that liver CYP4Fs are not the only players in metabolizing chlorpromazine. To study further the underlying mechanism, CYP4F5 gene was isolated, characterized, and the promoter region was defined. ^ Accumulating evidence showed that peroxisome proliferator-activated receptors (PPARs) play an active role in inflammation. To investigate the possible role of PPARα in regulating CYP4F expression by inflammation or by clofibrate treatment, the expressions of two new mouse 4F isoforms were analyzed in PPARα knockout mice upon LPS or clofibrate challenge. A novel induction of CYP4F15 by LPS and clofibrate was observed in kidney, and this effect is totally dependent on the presence of PPARα. Renal CYP4F16 expression was not affected by LPS or clofibrate in both (+/+) and (−/−) mice. In contrast, hepatic expressions of CYP4F15 and CYP4F16 were reduced significantly in (+/+) mice, but much less in (−/−) mice, suggesting that PPARα is partially responsible for this down-regulation. Clofibrate treatment reduced the expression of CYP4F16 in liver, but has no effect on CYP4F15 and PPARα does not have a role in hepatic CYP4F expression regulated by clofibrate. In general, CYP4Fs are regulated in an isoform-, tissue- and species-specific manner. ^ A human CYP4F isoform, CYP4F11, was isolated. The genomic structure was also solved by using database mining and bioinformatics tools. Localization of CYP4F11 to chromosome 19, 16 kb upstream of CYP4F2, suggests that human CYP4F genes may form a cluster on chromosome 19. This novel human 4F is highly expressed in liver, as well as in kidney, heart and skeletal muscle. Further study of the activity and gene regulation on CYP4F11 will provide us more insights into the physiological functions of CYP4F subfamily. ^

Role of ovarian cancer antigen CA125/MUC16 in development and cancer

Cancer antigen 125 (CA125) is a tumor antigen that is routinely used to monitor the disease progress and the outcome of treatment in ovarian cancer patients. Elevated serum levels of CA125 are detected in over 80% of epithelial ovarian cancer patients. CA125 is a high molecular weight (>1M Dalton) mucin-type glycoprotein encoded by the MUC16 gene on human chromosome 19. Although MUC16 has served as the best serum marker for monitoring growth of ovarian cancer, roles for MUC16 in normal physiology and ovarian cancer are largely unknown. To understand the biological functions of MUC16, I characterized a mouse Muc16 homolog on chromosome 9 by means of expression pattern profiling, phenotype analysis of Muc16 knockout mice, and in vitro and in vivo studies of Muc16 null transformed ovarian surface epithelial (OSE) cells. ^ The mouse Muc16 homolog shares a conserved genomic structure with human MUC16. In addition to being expressed in mouse ovarian cancer, mouse Muc16 mRNA and protein were expressed in the mesothelia covering the heart, lung, ovary, oviduct, spleen, testis, and uterus. The conserved genomic structure and expression pattern of mouse Muc16 to human MUC16 suggests that mouse Muc16 is the ortholog of human MUC16. To understand the biological functions of Muc16, I generated Muc16 knockout mice. Muc16 knockout mice were viable, fertile and normal by one year of age. However, between 18 and 24 months of age, Muc16 knockout mice developed various tissue abnormalities such as ovarian cysts and tumors of the liver and other peritoneal organs. To determine the role of MUC16 in ovarian cancer progression, I established Muc16 null transformed ovarian surface epithelial (OSE) cell lines, following the same method to develop mouse model of epithelial ovarian cancer (Orsulic et al., 2002). Loss of Muc16 did not affect cell morphology, cell proliferation rate, or tumorigenic potential. However, Muc16-null OSE cells showed decreased attachment to extracellular matrix proteins as well as to primary mouse peritoneal mesothelial cells. Peritoneal mesothelia are the most frequent implantation sites of ovarian cancer. Furthermore, a pilot transplantation assay suggests that Muc16 null transformed OSE cells formed less disseminated tumors in the peritoneal cavity compared to wild-type OSE cells. ^ In conclusion, these results demonstrate that MUC16 is not required for normal mouse development or reproduction, but plays important roles in tissue homeostasis, ovarian cancer cell adhesion and dissemination. This study provides the first in vivo evidence of the roles of MUC16 in development, as well as ovarian cancer progression and dissemination. These studies offer valuable insights into possible mechanisms of ovarian cancer development and potential molecular targets for ovarian cancer treatment. ^

Identification of a novel tumor suppressor gene on human chromosome 3p14-p12 involved in renal cell carcinoma

Nonpapillary renal cell carcinoma (RCC) is an adult cancer of the kidney which occurs both in familial and sporadic forms. The familial form of RCC is associated with translocations involving chromosome 3 with a breakpoint at 3p14-p13. Studies focused on sporadic RCC have shown two commonly deleted regions at 3p14.3-p13 and 3p21.3. In addition, a more distal region mapping to 3p26-p25 has been linked to the Von Hippel Lindau (VHL) disease gene. A large proportion of VHL patients develop RCC. The short arm of human chromosome 3 can, therefore, be dissected into three distinct regions which could encode tumor suppressor genes for RCC. Loss or inactivation of one or more of these loci may be an important step in the genesis of RCC.^ I have used the technique of microcell-mediated chromosome transfer to introduce an intact, normal human chromosome 3 and defined fragments of 3p, dominantly marked with pSV2neo, into the highly malignant RCC cell line SN12C.19. The introduction of chromosome 3 and of a centric fragment of 3p, encompassing 3p14-q11, into SN12C.19 resulted in dramatic suppression of tumor growth in nude mice. Another defined deletion hybrid contained the region 3p12-q24 of the introduced human chromosome and failed to suppress tumorigenicity. These data define the region 3p14-p12, the most proximal region of high frequency allele loss in sporadic RCC as well as the region containing the translocation breakpoint in familial RCC, to contain a novel tumor suppressor locus involved in RCC. We have designated this locus nonpapillary renal cell carcinoma-1 (NRC-1). Furthermore, we have functional evidence that NRC-1 controls the growth of RCC cells by inducing rapid cell death in vivo. ^