Chromosome 17 deletions and p53 gene mutations in colorectal carcinomas

Previous studies have demonstrated that allelic deletions of the short arm of chromosome 17 occur in over 75% of colorectal carcinomas. Twenty chromosome 17p markers were used to localize the common region of deletion in these tumors to a region contained within bands 17p12 to 17p13.3. This region contains the gene for the transformation-associated protein p53. Southern and Northern blot hybridization experiments provided no evidence for gross alterations of the p53 gene or surrounding sequences. As a more rigorous test of the possibility that p53 was a target of the deletions, the p53 coding regions from two tumors were analyzed; these two tumors, like most colorectal carcinomas, had allelic deletions of chromosome 17p and expressed considerable amounts of p53 messenger RNA from the remaining allele. The remaining p53 allele was mutated in both tumors, with an alanine substituted for valine at codon 143 of one tumor and a histidine substituted for arginine at codon 175 of the second tumor. Both mutations occurred in a highly conserved region of the p53 gene that was previously found to be mutated in murine p53 oncogenes. The data suggest that p53 gene mutations may be involved in colorectal neoplasia, perhaps through inactivation of a tumor suppressor function of the wild-type p53 gene.     

Concerted nonsyntenic allelic loss in human colorectal carcinoma

Familial polyposis coli (FPC) is caused by an autosomal dominant gene on chromosome 5, and it has been proposed that colorectal cancer in the general population arises from loss or inactivation of the FPC gene, analogous to recessive tumor genes in retinoblastoma and Wilms' tumor. Since allelic loss can be erroneously scored in nonhomogeneous samples, tumor cell populations were first microdissected from 24 colorectal carcinomas, an additional nine cancers were engrafted in nude mice, and nuclei were flow-sorted from an additional two. Of 31 cancers informative for chromosome 5 markers, only 6 (19%) showed loss of heterozygosity of chromosome 5 alleles, compared to 19 of 34 (56%) on chromosome 17, and 17 of 33 (52%) on chromosome 18. Therefore, it appears that (i) FPC is a true dominant for adenomatosis but not a common recessive gene for colon cancer; and (ii) simple Mendelian models involving loss of alleles at a single locus may be inappropriate for understanding common human solid tumors.     

A chicken transferrin gene in transgenic mice escapes X-chromosome inactivation

Mammalian X-chromosome inactivation involves a coordinate shutting down of physically linked genes. Several proposed models require the presence of specific sequences near genes to permit the spread of inactivation into these regions. If such models are correct, one might predict that heterologous genes transferred onto the X chromosome might lack the appropriate signal sequences and therefore escape inactivation. To determine whether a foreign gene inserted into the X chromosome is subject to inactivation, transgenic mice harboring 11 copies of the complete, 17-kilobase chicken transferrin gene on the X chromosome were used. Male mice hemizygous for this insert were bred with females bearing Searle's translocation, an X-chromosome rearrangement that is always active in heterozygous females (the unrearranged X chromosome is inactive). Female offspring bearing the Searle's translocation and the chicken transferrin gene had the same amount of chicken transferrin messenger RNA in liver as did transgenic male mice or transgenic female mice lacking the Searle's chromosome. This result shows that the inserted gene is not subject to X-chromosome inactivation and suggests that the inactivation process cannot spread over 187 kilobases of DNA in the absence of specific signal sequences required for inactivation.     

X-Chromosome inactivation in cloned mouse embryos

To study whether cloning resets the epigenetic differences between the two X chromosomes of a somatic female nucleus, we monitored X inactivation in cloned mouse embryos. Both X chromosomes were active during cleavage of cloned embryos, followed by random X inactivation in the embryo proper. In the trophectoderm (TE), X inactivation was nonrandom with the inactivated X of the somatic donor being chosen for inactivation. When female embryonic stem cells with two active X chromosomes were used as donors, random X inactivation was seen in the TE and embryo. These results demonstrate that epigenetic marks can be removed and reestablished on either X chromosome during cloning. Our results also suggest that the epigenetic marks imposed on the X chromosomes during gametogenesis, responsible for normal imprinted X inactivation in the TE, are functionally equivalent to the marks imposed on the chromosomes during somatic X inactivation.     

An X chromosome gene, WTX, is commonly inactivated in Wilms tumor

Wilms tumor is a pediatric kidney cancer associated with inactivation of the WT1 tumor-suppressor gene in 5 to 10% of cases. Using a high-resolution screen for DNA copy-number alterations in Wilms tumor, we identified somatic deletions targeting a previously uncharacterized gene on the X chromosome. This gene, which we call WTX, is inactivated in approximately one-third of Wilms tumors (15 of 51 tumors). Tumors with mutations in WTX lack WT1 mutations, and both genes share a restricted temporal and spatial expression pattern in normal renal precursors. In contrast to biallelic inactivation of autosomal tumor-suppressor genes, WTX is inactivated by a monoallelic "single-hit" event targeting the single X chromosome in tumors from males and the active X chromosome in tumors from females.     

Reduction to homozygosity of genes on chromosome 11 in human breast neoplasia

The somatic loss of heterozygosity for normal alleles occurring in human tumors has suggested the presence of recessive oncogenes. The results presented here demonstrate a loss of heterozygosity of several genes on chromosome 11 in primary breast tumors. Restriction fragment length polymorphism analysis of these DNAs further suggests that the most frequent loss of sequences in breast tumors occurs between the beta-globin and parathyroid hormone loci on the short arm of chromosome 11. The loss of heterozygosity for chromosome 11 loci has a significant association with tumors that lack estrogen and progesterone receptors, grade III tumors, and distal metastasis.     

Identification of a gene located at chromosome 5q21 that is mutated in colorectal cancers.

Recent studies have suggested the existence of a tumor suppressor gene located at chromosome region 5q21. DNA probes from this region were used to study a panel of sporadic colorectal carcinomas. One of these probes, cosmid 5.71, detected a somatically rearranged restriction fragment in the DNA from a single tumor. Further analysis of the 5.71 cosmid revealed two regions that were highly conserved in rodent DNA. These sequences were used to identify a gene, MCC (mutated in colorectal cancer), which encodes an 829-amino acid protein with a short region of similarity to the G protein-coupled m3 muscarinic acetylcholine receptor. The rearrangement in the tumor disrupted the coding region of the MCC gene. Moreover, two colorectal tumors were found with somatically acquired point mutations in MCC that resulted in amino acid substitutions. MCC is thus a candidate for the putative colorectal tumor suppressor gene located at 5q21. Further studies will be required to determine whether the gene is mutated in other sporadic tumors or in the germ line of patients with an inherited predisposition to colonic tumorigenesis.     

结直肠癌中CXCR7、VEGF、Ki-67和PTEN表达

目的探讨结直肠癌中CXCR7、VEGF、Ki-67和PTEN表达情况及意义。方法采用免疫组化Envision法检测CXCR7、VEGF、Ki-67和PTEN在60例结直肠癌、30例结直肠腺瘤、10例正常结直肠黏膜组织中表达。结果 CXCR7、VEGF、Ki-67在结直肠癌中表达明显高于腺瘤和正常黏膜（P〈0.01）,而PTEN表达降低（P〈0.01）。CXCR7、VEGF、Ki-67表达与肿瘤直径、淋巴结转移有关（P〈0.01或0.05）,而PTEN表达与肿瘤分化程度有关（P〈0.05）。结直肠癌中CXCR7表达与VEGF和Ki-67表达有关（P〈0.01或0.05）。结论结直肠癌中CXCR7、VEGF高表达参与肿瘤增殖和淋巴结转移,而PTEN低表达可能与肿瘤低分化有关。     

Autosomal dominant mutations affecting X inactivation choice in the mouse

X chromosome inactivation is the silencing mechanism eutherian mammals use to equalize the expression of X-linked genes between males and females early in embryonic development. In the mouse, genetic control of inactivation requires elements within the X inactivation center (Xic) on the X chromosome that influence the choice of which X chromosome is to be inactivated in individual cells. It has long been posited that unidentified autosomal factors are essential to the process. We have used chemical mutagenesis in the mouse to identify specific factors involved in X inactivation and report two genetically distinct autosomal mutations with dominant effects on X chromosome choice early in embryogenesis.     

Gene for T-cell growth factor: location on human chromosome 4q and feline chromosome B1

T-cell growth factor (TCGF) or interleukin-2 (IL-2), an immunoregulatory lymphokine, is produced by lectin- or antigen-activated mature T lymphocytes and in a constitutive manner by certain T-cell lymphoma cell lines. By means of a molecular clone of human TCGF and DNA extracted from a panel of somatic cell hybrids (rodent cells X normal human lymphocytes), the TCGF structural gene was identified on human chromosome 4. In situ hybridization of the TCGF clone to human chromosomes resulted in significant labeling of the midportion of the long arm of chromosome 4, indicating that the TCGF gene was located at band q26-28. Genomic DNA from a panel of hybrids prepared with HUT-102 B2 cells was examined with the same molecular clone. In this clone of cells, which produces human T-cell leukemia virus, the TCGF gene was also located on chromosome 4 and was apparently not rearranged. The homologous TCGF locus in the domestic cat was assigned to chromosome B1 by using a somatic cell hybrid panel that segregates cat chromosomes. Linkage studies as well as high-resolution G-trypsin banding indicate that this feline chromosome is partially homologous to human chromosome 4.     

Reactivation of an inactive human X chromosome: evidence for X inactivation by DNA methylation

A mouse-human somatic cell hybrid clone, deficient in hypoxanthine-guanine phosphoribosyltransferase (HPRT) and containing a structurally normal inactive human X chromosome, was isolated. The hybrid cells were treated with 5-azacytidine and tested for the reactivation and expression of human X-linked genes. The frequency of HPRT-positives clones after 5-azacytidine treatment was 1000-fold greater than that observed in untreated hybrid cells. Fourteen independent HPRT-positive clones were isolated and analyzed for the expression of human X markers. Isoelectric focusing showed that the HPRT expressed in these clones is human. One of the 14 clones expressed human glucose-6-phosphate dehydrogenase and another expressed human phosphoglycerate kinase. Since 5-azacytidine treatment results in hypomethylation of DNA, DNA methylation may be a mechanism of human X chromosome inactivation.     

Translocation and rearrangement of myeloperoxidase gene in acute promyelocytic leukemia

Acute promyelocytic leukemia (subtype M3) is characterized by malignant promyelocytes exhibiting an abundance of abnormally large or aberrant primary granules. Myeloperoxidase (MPO) activity of these azurophilic granules, as assessed by cytochemical staining, is unusually intense. In addition, M3 is universally associated with a chromosomal translocation, t(15;17)(q22;q11.2). In this report, the MPO gene was localized to human chromosome 17 (q12-q21), the region of the breakpoint on chromosome 17 in the t(15;17), by somatic cell hybrid analysis and in situ chromosomal hybridization. By means of MPO complementary DNA clones for in situ hybridization and Southern blot analysis, the effect of this specific translocation on the MPO gene was examined. In all cases of M3 examined, MPO is translocated to chromosome 15. Genomic blot analyses indicate rearrangement of MPO in leukemia cells of two of four cases examined. These findings suggest that MPO may be pivotal in the pathogenesis of acute promyelocytic leukemia.     

Epigenetic dynamics of imprinted X inactivation during early mouse development

The initiation of X-chromosome inactivation is thought to be tightly correlated with early differentiation events during mouse development. Here, we show that although initially active, the paternal X chromosome undergoes imprinted inactivation from the cleavage stages, well before cellular differentiation. A reversal of the inactive state, with a loss of epigenetic marks such as histone modifications and polycomb proteins, subsequently occurs in cells of the inner cell mass (ICM), which give rise to the embryo-proper in which random X inactivation is known to occur. This reveals the remarkable plasticity of the X-inactivation process during preimplantation development and underlines the importance of the ICM in global reprogramming of epigenetic marks in the early embryo.     

Gene for alpha-chain of human T-cell receptor: location on chromosome 14 region involved in T-cell neoplasms

A human complementary DNA clone specific for the alpha-chain of the T-cell receptor and a panel of rodent X human somatic cell hybrids were used to map the alpha-chain gene to human chromosome 14 in a region proximal to the immunoglobulin heavy chain locus. Analysis by means of in situ hybridization of human metaphase chromosomes served to further localize the alpha-chain gene to region 14q11q12, which is consistently involved in translocations and inversions detectable in human T-cell leukemias and lymphomas. Thus, the locus for the alpha-chain T-cell receptor may participate in oncogene activation in T-cell tumors.     

非编码RNA Xist调节X染色体失活的分子机制

非编码KNAXist介导的X染色体失活是表观遗传研究的一个典范,该系统能使一整条染色体变为异染状态。从Xist与X染色体计数、失活的选择、失活的起始和失活的维持等方面综述了其分子机制。     

A phosphatase associated with metastasis of colorectal cancer

To gain insights into the molecular basis for metastasis, we compared the global gene expression profile of metastatic colorectal cancer with that of primary cancers, benign colorectal tumors, and normal colorectal epithelium. Among the genes identified, the PRL-3 protein tyrosine phosphatase gene was of particular interest. It was expressed at high levels in each of 18 cancer metastases studied but at lower levels in nonmetastatic tumors and normal colorectal epithelium. In 3 of 12 metastases examined, multiple copies of the PRL-3 gene were found within a small amplicon located at chromosome 8q24.3. These data suggest that the PRL-3 gene is important for colorectal cancer metastasis and provide a new therapeutic target for these intractable lesions.     

Allelotype of colorectal carcinomas

To examine the extent and variation of allelic loss in a common adult tumor, polymorphic DNA markers were studied from every nonacrocentric autosomal arm in 56 paired colorectal carcinoma and adjacent normal colonic mucosa specimens. This analysis was termed an allelotype, in analogy with a karyotype. Three major conclusions were drawn from this analysis: (i) Allelic deletions were remarkably common; one of the alleles of each polymorphic marker tested was lost in at least some tumors, and some tumors lost more than half of their parental alleles. (ii) In addition to allelic deletions, new DNA fragments not present in normal tissue were identified in five carcinomas; these new fragments contained repeated sequences of the variable number of tandem repeat type. (iii) Patients with more than the median percentage of allelic deletions had a considerably worse prognosis than did the other patients, although the size and stage of the primary tumors were very similar in the two groups. In addition to its implications concerning the genetic events underlying tumorigenesis, tumor allelotype may provide a molecular tool for improved estimation of prognosis in patients with colorectal cancer.     

Fragile X genotype characterized by an unstable region of DNA

DNA sequences have been located at the fragile X site by in situ hybridization and by the mapping of breakpoints in two somatic cell hybrids that were constructed to break at the fragile site. These hybrids were found to have breakpoints in a common 5-kilobase Eco RI restriction fragment. When this fragment was used as a probe on the chromosomal DNA of normal and fragile X genotype individuals, alterations in the mobility of the sequences detected by the probe were found only in fragile X genotype DNA. These sequences were of an increased size in all fragile X individuals and varied within families, indicating that the region was unstable. This probe provides a means with which to analyze fragile X pedigrees and is a diagnostic reagent for the fragile X genotype.     

The fragile X site in somatic cell hybrids: an approach for molecular cloning of fragile sites

Fragile X syndrome is a common form of mental retardation associated with a fragile site on the human X chromosome. Although fragility at this site is usually evident as a nonstaining chromatid gap, it remains unclear whether or not actual chromosomal breakage occurs. By means of somatic cell hybrids containing either a normal human X or a fragile X chromosome and utilizing two genes that flank the fragile site as markers of chromosome integrity, segregation of these markers was shown to be more frequent if they encompass the fragile site under appropriate culture conditions. Hybrid cells that reveal marker segregation were found to contain rearranged X chromosomes involving the region at or near the fragile site, thus demonstrating true chromosomal breakage within this area. Two independent translocation chromosomes were identified involving a rodent chromosome joined to the human X at the location of the fragile site. DNA analysis of closely linked, flanking loci was consistent with the position of the breakpoint being at or very near the fragile X site. Fragility at the translocation junctions was observed in both hybrids, but at significantly lower frequencies than that seen in the intact X of the parental hybrid. This observation suggests that the human portion of the junctional DNA may contain part of a repeated fragility sequence. Since the translocation junctions join heterologous DNA, the molecular cloning of the fragile X sequence should now be possible.