ABSTRACT
The human genome contains hundreds of large, structurally diverse blocks that are insufficiently represented in the reference genome and are thus not amenable to genomic analyses. Structural diversity in the human population suggests that these blocks are unstable in the germline; however, whether or not these blocks are also unstable in the cancer genome remains elusive. Here we report that the 500 kb block called KRTAP_region_1 (KRTAP-1) on 17q12-21 recurrently demarcates the amplicon of the ERBB2 (HER2) oncogene in breast tumors. KRTAP-1 carries numerous tandemly-duplicated segments that exhibit diversity within the human population. We evaluated the fragility of the block by cytogenetically measuring the distances between the flanking regions and found that spontaneous distance outliers (i.e DNA breaks) appear more frequently at KRTAP-1 than at the representative common fragile site (CFS) FRA16D. Unlike CFSs, KRTAP-1 is not sensitive to aphidicolin. The exonuclease activity of DNA repair protein Mre11 protects KRTAP-1 from breaks, whereas CtIP does not. Breaks at KRTAP-1 lead to the palindromic duplication of the ERBB2 locus and trigger Breakage-Fusion-Bridge cycles. Our results indicate that an insufficiently investigated area of the human genome is fragile and could play a crucial role in cancer genome evolution.
Subject(s)
Breast Neoplasms/genetics , Chromosome Fragile Sites/genetics , DNA Repair , Gene Amplification , Gene Duplication/genetics , Genes, erbB-2 , Keratins, Hair-Specific/physiology , Aphidicolin/pharmacology , Breast/metabolism , Breast Neoplasms/metabolism , Cells, Cultured , Chromosomal Instability , DNA Breaks , DNA Copy Number Variations , DNA, Neoplasm/genetics , Epithelial Cells/metabolism , Female , Genetic Variation , Genomic Instability , Humans , MRE11 Homologue Protein/physiology , Neoplasm Proteins/physiology , Whole Genome SequencingABSTRACT
AIM: To determine the role of Sonic hedgehog (Shh) pathway in colorectal adenocarcinomas through analysis of the expression of Shh pathway-related molecules, Shh, Ptch1, hedgehog-interacting protein (Hip), Gli1, Gli3 and PDGFRalpha. METHODS: Expression of Shh in 25 colorectal adeno-carcinomas was detected by RT-PCR, in situ hybridization and immunohistochemistry. Expression of Ptch1 was observed by in situ hybridization and immunohistochemistry. Expression of Hip, Gli1, Gli3 and PDGFRalpha was analyzed by in situ hybridization. RESULTS: Expression of cytokeratin AE1/AE3 was observed in the cytoplasm of colorectal crypts. Members of the Hh signaling pathway were expressed in colorectal epithelium. Shh was expressed in cytoplasm of dysplastic epithelial cells, while expression of Ptch1, Hip and Gli1 were mainly detected in the malignant crypts of adenocarcinomas. In contrast, PDGFRalpha was expressed highly in aberrant crypts and moderately in the stroma. Expression of Gli3 could not be detected in colorectal adenocarcinomas. CONCLUSION: These data suggest that Shh-Ptch1-Gli1 signaling pathway may play a role in the progression of colorectal tumor.
Subject(s)
Adenocarcinoma/physiopathology , Colorectal Neoplasms/physiopathology , Hedgehog Proteins/physiology , Oncogene Proteins/physiology , Signal Transduction/physiology , Trans-Activators/physiology , Adenocarcinoma/genetics , Adenocarcinoma/pathology , Carrier Proteins/genetics , Carrier Proteins/physiology , Colorectal Neoplasms/genetics , Colorectal Neoplasms/pathology , Gene Expression Regulation, Neoplastic , Hedgehog Proteins/genetics , Humans , Keratins, Hair-Specific/genetics , Keratins, Hair-Specific/physiology , Kruppel-Like Transcription Factors/genetics , Kruppel-Like Transcription Factors/physiology , Membrane Glycoproteins/genetics , Membrane Glycoproteins/physiology , Nerve Tissue Proteins/genetics , Nerve Tissue Proteins/physiology , Oncogene Proteins/genetics , Patched Receptors , Patched-1 Receptor , RNA, Messenger/genetics , RNA, Messenger/physiology , Receptor, Platelet-Derived Growth Factor alpha/genetics , Receptor, Platelet-Derived Growth Factor alpha/physiology , Receptors, Cell Surface/genetics , Receptors, Cell Surface/physiology , Trans-Activators/genetics , Zinc Finger Protein GLI1 , Zinc Finger Protein Gli3ABSTRACT
Hair follicles cycle between stages of growth (anagen) and metabolic quiescence (telogen) throughout life. In mature follicles, transition from telogen back into anagen involves the activation, proliferation, and differentiation of epithelial stem cells located in the bulge, a specialization of the outer root sheath. Recent studies identified keratin 6a (K6a) transcripts as enriched in bulge epithelial stem cells in mouse skin. We used messenger RNA probes, antibodies, a LacZ reporter mouse model, and whole-mount staining assays to investigate the regulation of mK6a during mouse postnatal hair cycling, and compare it to mK75, a companion layer (Cl) marker. We find that mK75 regulation parallels that of inner root sheath (IRS) markers, with expression onset at anagen IIIa above the new hair bulb and subsequent spreading towards the bulge. Although also occurring in the Cl, mK6a expression begins at anagen IIIb in differentiating cells located proximal to the bulge, and subsequently spreads towards the hair bulb. mK6a and mK75 thus exhibit temporally distinct, and spatially opposed, expression patterns in the Cl during postnatal anagen. These findings provide novel insight into the morphogenesis and properties of the Cl, and raise the distinct possibility that it is an integral part of the IRS compartment.