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Differential expression of chaperonin containing T-complex polypeptide (CCT) subunits during fetal and adult skin wound healing.

ABSTRACT: Integumentary wound healing in early fetal life is regenerative and proceeds without scar formation. Expressomic analysis of this phenomenon by differential display has previously determined that the eta subunit of the cytosolic chaperonin containing T-complex polypeptide (CCT) is downregulated in the healing fetal wound milieu. We now report that no other CCT subunit shares this distinct pattern of gene regulation as determined by limiting dilution reverse transcriptase polymerase chain reaction (RT-PCR); all seven of the remaining CCT subunits demonstrate no change in messenger RNA (mRNA) expression in healing fetal wounds compared to unwounded control tissue. The alpha subunit, however, did evidence reduced message levels in healing adult wound tissue. We herein report on the cloning and sequence of the complementary DNA (cDNA) for rabbit CCT-alpha and confirm its wound specific decrease in adult tissues through quantitative real-time RT-PCR assay. We also confirm that quantitative evaluation of CCT-alpha and CCT-zeta mRNA expression shows no change in healing fetal wounds.

SUBMITTER: Satish L

PROVIDER: S-EPMC2673928 | biostudies-other | 2008 Dec

REPOSITORIES: biostudies-other

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Project description:Chaperonin containing TCP1 (CCT/TRiC) is a multi-subunit protein folding complex that enables the cancer phenotype to emerge from the mutational landscape that drives oncogenesis. We and others linked increased expression of CCT subunits to advanced tumor stage and invasiveness that inversely correlates with cancer patient outcomes. In this study, we examined the expression of the second CCT subunit, CCT2, using genomic databases of adult and pediatric tumors and normal tissues, and found that it was highly expressed in pediatric cancers, showing a significant difference compared to normal tissues. Histologic staining confirmed that CCT subunits are highly expressed in tumor tissues, which was exemplified in neuroblastoma. Using two neuroblastoma cells, MYCN-amplified, IMR-32 cells, and non-amplified, SK-N-AS cells, we assessed baseline levels for CCT subunits and found expressions comparable to the highly invasive triple-negative breast cancer (TNBC) cell line, MDA-MB-231. Exogenous expression of CCT2 in both SK-N-AS and IMR-32 cells resulted in morphological changes, such as larger cell size and increased adherence, with significant increases in the CCT substrates, actin, and tubulin, as well as increased migration. Depletion of CCT2 reversed these effects and reduced cell viability. We evaluated CCT as a therapeutic target in IMR-32 cells by testing a novel peptide CCT inhibitor, CT20p. Treatment with CT20p induced cell death in these neuroblastoma cells. The use of CCT2 as a biological indicator for detection of neuroblastoma cells shed in blood was examined by spiking IMR-32 cells into human blood and using an anti-CCT2 antibody for the identification of spiked cancer cells with the CellSearch system. Results showed that using CCT2 for the detection of neuroblastoma cells in blood was more effective than the conventional approach of using epithelial markers like cytokeratins. CCT2 plays an essential role in promoting the invasive capacity of neuroblastoma cells and thus offers the potential to act as a molecular target in the development of novel therapeutics and diagnostics for pediatric cancers.

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Differential expression of chaperonin containing T-complex polypeptide (CCT) subunits during fetal and adult skin wound healing.

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