Limits...
KIAA0101 (OEACT-1), an expressionally down-regulated and growth-inhibitory gene in human hepatocellular carcinoma.

Guo M, Li J, Wan D, Gu J - BMC Cancer (2006)

Bottom Line: The in vitro cell growth curve was used for examining the effect of over-expression of KIAA0101 in HCC cells.Western blot showed KIAA0101 protein expression was down-regulated in HCC tissues as compared with their counterpart non-cancerous liver tissues in 25 out of 30 cases.This gene could inhibit the HCC cell growth in vitro and presumably by its blocking effect on cell cycle.

View Article: PubMed Central - HTML - PubMed

Affiliation: Shanghai Medical College, Fudan University, Shanghai, China. guomingleicn@yahoo.com.cn

ABSTRACT

Background: Our previous cDNA array results indicated KIAA0101 as one of the differentially expressed genes in human hepatocellular carcinoma (HCC) as compared with non-cancerous liver. However, it is necessary to study its expression at protein level in HCC and its biological function for HCC cell growth.

Method: Western blot and tissue array were performed to compare KIAA0101 protein expression level in paired human HCC and non-cancerous liver tissues from the same patients. Investigation of its subcellular localization was done by using dual fluorescence image examination and enriched mitochondrial protein Western blot analysis. The in vitro cell growth curve was used for examining the effect of over-expression of KIAA0101 in HCC cells. FACS was used to analyze the cell cycle pattern in KIAA0101 expression positive (+) and negative (-) cell populations isolated by the pMACSKKII system after KIAA0101 cDNA transfection.

Results: Western blot showed KIAA0101 protein expression was down-regulated in HCC tissues as compared with their counterpart non-cancerous liver tissues in 25 out of 30 cases. Tissue array also demonstrated the same pattern in 161 paired samples. KIAA0101 was predominantly localized in mitochondria and partially in nuclei. KIAA0101 cDNA transfection could inhibit the HCC cell growth in vitro. In cell cycle analysis, it could arrest cells at the G1 to S phase transition.

Conclusion: KIAA0101 protein expression was down-regulated in HCC. This gene could inhibit the HCC cell growth in vitro and presumably by its blocking effect on cell cycle.

Show MeSH

Related in: MedlinePlus

Western blot for KIAA0101 protein in mitochondrial-enriched protein fraction and total cell proteins (1 and 2 were mitochondrial enriched proteins from SMMC 7721 and HepG2 respectively; 3 and 4 were total cell protein from SMMC 7721 and HepG2 respectively).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC1483895&req=5

Figure 5: Western blot for KIAA0101 protein in mitochondrial-enriched protein fraction and total cell proteins (1 and 2 were mitochondrial enriched proteins from SMMC 7721 and HepG2 respectively; 3 and 4 were total cell protein from SMMC 7721 and HepG2 respectively).

Mentions: The bioinformatics has implicated that KIAA0101 was possibly localized in mitochondrial. To verify this prediction, we have performed immuno-fluorescence survey on SMMC7721 cells by using the polyclonal rabbit antibody. KIAA0101 immuno-fluorescence was exhibited in a speckled pattern in the cytoplasm. The distribution of KIAA0101 immuno-fluorescence appeared to be similar to the distribution of mitochondria. In dual-labeling experiments, KIAA0101 immuno-fluorescence partially coincided with the image generated by a labeled monoclonal antibody that recognized cytochrome oxidase subunit I, an inner mitochondrial membrane protein. The partially co-localization of KIAA0101 with mitochondrial proteins suggested that KIAA0101 was localized predominantly in mitochondria (Fig 4). Meanwhile, we used the specific mouse anti-mitochondrial (Neomarkers) to enrich the mitochondrial proteins from total cell proteins preparation. After precipitated by the protein A/G beads, the enriched mitochondrial proteins were subjected to Western blot with KIAA0101 rabbit antibody. Fig 5 showed that there were strong signal in lanes loaded by enriched mitochondrial proteins; however, there were only very weak signal in the un-sorted total cell proteins. From these two different experiments, we could conclude that the KIAA0101 protein was predominantly located in the mitochondria of human HCC SMMC 7721 cells.


KIAA0101 (OEACT-1), an expressionally down-regulated and growth-inhibitory gene in human hepatocellular carcinoma.

Guo M, Li J, Wan D, Gu J - BMC Cancer (2006)

Western blot for KIAA0101 protein in mitochondrial-enriched protein fraction and total cell proteins (1 and 2 were mitochondrial enriched proteins from SMMC 7721 and HepG2 respectively; 3 and 4 were total cell protein from SMMC 7721 and HepG2 respectively).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC1483895&req=5

Figure 5: Western blot for KIAA0101 protein in mitochondrial-enriched protein fraction and total cell proteins (1 and 2 were mitochondrial enriched proteins from SMMC 7721 and HepG2 respectively; 3 and 4 were total cell protein from SMMC 7721 and HepG2 respectively).
Mentions: The bioinformatics has implicated that KIAA0101 was possibly localized in mitochondrial. To verify this prediction, we have performed immuno-fluorescence survey on SMMC7721 cells by using the polyclonal rabbit antibody. KIAA0101 immuno-fluorescence was exhibited in a speckled pattern in the cytoplasm. The distribution of KIAA0101 immuno-fluorescence appeared to be similar to the distribution of mitochondria. In dual-labeling experiments, KIAA0101 immuno-fluorescence partially coincided with the image generated by a labeled monoclonal antibody that recognized cytochrome oxidase subunit I, an inner mitochondrial membrane protein. The partially co-localization of KIAA0101 with mitochondrial proteins suggested that KIAA0101 was localized predominantly in mitochondria (Fig 4). Meanwhile, we used the specific mouse anti-mitochondrial (Neomarkers) to enrich the mitochondrial proteins from total cell proteins preparation. After precipitated by the protein A/G beads, the enriched mitochondrial proteins were subjected to Western blot with KIAA0101 rabbit antibody. Fig 5 showed that there were strong signal in lanes loaded by enriched mitochondrial proteins; however, there were only very weak signal in the un-sorted total cell proteins. From these two different experiments, we could conclude that the KIAA0101 protein was predominantly located in the mitochondria of human HCC SMMC 7721 cells.

Bottom Line: The in vitro cell growth curve was used for examining the effect of over-expression of KIAA0101 in HCC cells.Western blot showed KIAA0101 protein expression was down-regulated in HCC tissues as compared with their counterpart non-cancerous liver tissues in 25 out of 30 cases.This gene could inhibit the HCC cell growth in vitro and presumably by its blocking effect on cell cycle.

View Article: PubMed Central - HTML - PubMed

Affiliation: Shanghai Medical College, Fudan University, Shanghai, China. guomingleicn@yahoo.com.cn

ABSTRACT

Background: Our previous cDNA array results indicated KIAA0101 as one of the differentially expressed genes in human hepatocellular carcinoma (HCC) as compared with non-cancerous liver. However, it is necessary to study its expression at protein level in HCC and its biological function for HCC cell growth.

Method: Western blot and tissue array were performed to compare KIAA0101 protein expression level in paired human HCC and non-cancerous liver tissues from the same patients. Investigation of its subcellular localization was done by using dual fluorescence image examination and enriched mitochondrial protein Western blot analysis. The in vitro cell growth curve was used for examining the effect of over-expression of KIAA0101 in HCC cells. FACS was used to analyze the cell cycle pattern in KIAA0101 expression positive (+) and negative (-) cell populations isolated by the pMACSKKII system after KIAA0101 cDNA transfection.

Results: Western blot showed KIAA0101 protein expression was down-regulated in HCC tissues as compared with their counterpart non-cancerous liver tissues in 25 out of 30 cases. Tissue array also demonstrated the same pattern in 161 paired samples. KIAA0101 was predominantly localized in mitochondria and partially in nuclei. KIAA0101 cDNA transfection could inhibit the HCC cell growth in vitro. In cell cycle analysis, it could arrest cells at the G1 to S phase transition.

Conclusion: KIAA0101 protein expression was down-regulated in HCC. This gene could inhibit the HCC cell growth in vitro and presumably by its blocking effect on cell cycle.

Show MeSH
Related in: MedlinePlus