Abstract
Mutations in the von Hippel-Lindau (VHL) gene are involved in the VHL family cancer syndrome and sporadic renal cell carcinoma. Previous studies indicated that VHL-induced growth arrest required high cell density and growth on extracellular matrix. In the present study, VHL protein (pVHL) levels were observed to be dramatically increased in cells grown to high cell density compared to cells grown at low cell density. Reverse transcription-polymerase chain reaction and Northern blot analysis indicated that VHL mRNA levels were equivalent in sparse and dense cells. The pVHL was rapidly degraded when cell-cell contact was disturbed by trypsinization or EDTA release. Treatment of cells with a proteasome inhibitor blocked the degradation of pVHL. Using a set of VHL deletions fused to GFP, a cell density-dependent region (CDDR) was identified and localized to the c-terminus of pVHL. In addition, other members of the VBC protein complex also showed a cell density-dependent regulation similar to pVHL. Cell density regulation of VHL did not require elongin binding and density-dependent regulation of other VBC components was not dependent on pVHL. In addition, hypoxia inducible factor-2α protein levels were elevated in sparse cells with low levels of pVHL and reduced or absent in confluent cells containing abundant VHL. These results indicate that pVHL levels and thus function are tightly regulated by cell-cell signaling. In addition, care must be taken when interpreting studies of VHL function and subcellular localization of cells grown at subconfluent conditions.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 5270-5280 |
| Number of pages | 11 |
| Journal | Oncogene |
| Volume | 22 |
| Issue number | 34 |
| DOIs | |
| State | Published - Aug 14 2003 |
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Keywords
- Cell density
- Cell-cell contact
- VHL
ASJC Scopus subject areas
- Molecular Biology
- Cancer Research
- Genetics
Cite this
Von Hippel-Lindau protein complex is regulated by cell density. / Mohan, Sankar; Burk, Robert D.
In: Oncogene, Vol. 22, No. 34, 14.08.2003, p. 5270-5280.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Von Hippel-Lindau protein complex is regulated by cell density
AU - Mohan, Sankar
AU - Burk, Robert D.
PY - 2003/8/14
Y1 - 2003/8/14
N2 - Mutations in the von Hippel-Lindau (VHL) gene are involved in the VHL family cancer syndrome and sporadic renal cell carcinoma. Previous studies indicated that VHL-induced growth arrest required high cell density and growth on extracellular matrix. In the present study, VHL protein (pVHL) levels were observed to be dramatically increased in cells grown to high cell density compared to cells grown at low cell density. Reverse transcription-polymerase chain reaction and Northern blot analysis indicated that VHL mRNA levels were equivalent in sparse and dense cells. The pVHL was rapidly degraded when cell-cell contact was disturbed by trypsinization or EDTA release. Treatment of cells with a proteasome inhibitor blocked the degradation of pVHL. Using a set of VHL deletions fused to GFP, a cell density-dependent region (CDDR) was identified and localized to the c-terminus of pVHL. In addition, other members of the VBC protein complex also showed a cell density-dependent regulation similar to pVHL. Cell density regulation of VHL did not require elongin binding and density-dependent regulation of other VBC components was not dependent on pVHL. In addition, hypoxia inducible factor-2α protein levels were elevated in sparse cells with low levels of pVHL and reduced or absent in confluent cells containing abundant VHL. These results indicate that pVHL levels and thus function are tightly regulated by cell-cell signaling. In addition, care must be taken when interpreting studies of VHL function and subcellular localization of cells grown at subconfluent conditions.
AB - Mutations in the von Hippel-Lindau (VHL) gene are involved in the VHL family cancer syndrome and sporadic renal cell carcinoma. Previous studies indicated that VHL-induced growth arrest required high cell density and growth on extracellular matrix. In the present study, VHL protein (pVHL) levels were observed to be dramatically increased in cells grown to high cell density compared to cells grown at low cell density. Reverse transcription-polymerase chain reaction and Northern blot analysis indicated that VHL mRNA levels were equivalent in sparse and dense cells. The pVHL was rapidly degraded when cell-cell contact was disturbed by trypsinization or EDTA release. Treatment of cells with a proteasome inhibitor blocked the degradation of pVHL. Using a set of VHL deletions fused to GFP, a cell density-dependent region (CDDR) was identified and localized to the c-terminus of pVHL. In addition, other members of the VBC protein complex also showed a cell density-dependent regulation similar to pVHL. Cell density regulation of VHL did not require elongin binding and density-dependent regulation of other VBC components was not dependent on pVHL. In addition, hypoxia inducible factor-2α protein levels were elevated in sparse cells with low levels of pVHL and reduced or absent in confluent cells containing abundant VHL. These results indicate that pVHL levels and thus function are tightly regulated by cell-cell signaling. In addition, care must be taken when interpreting studies of VHL function and subcellular localization of cells grown at subconfluent conditions.
KW - Cell density
KW - Cell-cell contact
KW - VHL
UR - http://www.scopus.com/inward/record.url?scp=0041353455&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0041353455&partnerID=8YFLogxK
U2 - 10.1038/sj.onc.1206592
DO - 10.1038/sj.onc.1206592
M3 - Article
VL - 22
SP - 5270
EP - 5280
JO - Oncogene
JF - Oncogene
SN - 0950-9232
IS - 34
ER -