Cyclin I activates Cdk5 and regulates expression of Bcl-2 and Bcl-XL in postmitotic mouse cells
Paul T. Brinkkoetter, … , James M. Roberts, Stuart J. Shankland
Paul T. Brinkkoetter, … , James M. Roberts, Stuart J. Shankland
Published September 1, 2009
Citation Information: J Clin Invest. 2009;119(10):3089-3101. https://doi.org/10.1172/JCI37978.
View: Text | PDF
Research Article Nephrology

Cyclin I activates Cdk5 and regulates expression of Bcl-2 and Bcl-XL in postmitotic mouse cells

Abstract

Cyclin I is an atypical cyclin because it is most abundant in postmitotic cells. We previously showed that cyclin I does not regulate proliferation, but rather controls survival of podocytes, terminally differentiated epithelial cells that are essential for the structural and functional integrity of kidney glomeruli. Here, we investigated the mechanism by which cyclin I safeguards against apoptosis and found that cyclin I bound and activated cyclin-dependent kinase 5 (Cdk5) in isolated mouse podocytes and neurons. Cdk5 activity was reduced in glomeruli and brain lysates from cyclin I–deficient mice, and inhibition of Cdk5 increased in vitro the susceptibility to apoptosis in response to cellular damage. In addition, levels of the prosurvival proteins Bcl-2 and Bcl-XL were reduced in podocytes and neurons from cyclin I–deficient mice, and restoration of Bcl-2 or Bcl-XL expression prevented injury-induced apoptosis. Furthermore, we found that levels of phosphorylated MEK1/2 and ERK1/2 were decreased in cyclin I–deficient podocytes and that inhibition of MEK1/2 restored Bcl2 and Bcl-XL protein levels. Of interest, this pathway was also defective in mice with experimental glomerulonephritis. Taken together, these data suggest that a cyclin I–Cdk5 complex forms a critical antiapoptotic factor in terminally differentiated cells that functions via MAPK signaling to modulate levels of the prosurvival proteins Bcl-2 and Bcl-XL.

Authors

Paul T. Brinkkoetter, Paul Olivier, Jimmy S. Wu, Scott Henderson, Ronald D. Krofft, Jeffrey W. Pippin, David Hockenbery, James M. Roberts, Stuart J. Shankland

×

Figure 4

Cyclin I–Cdk5 kinase activity is required to protect podocytes from apoptosis.

Options: View larger image (or click on image) Download as PowerPoint
Cyclin I–Cdk5 kinase activity is required to protect podocytes from apop...
(A) Apoptosis was quantitated by Hoechst 33342 staining in podocytes under nonstressed conditions (baseline) and 6 hours after UV-C irradiation (25 J/m2). UV-C–induced apoptosis was increased 2- to 3-fold in WT podocytes transfected with siRNA-targeting Cdk5 (lane 3) compared with cells transfected with control siRNA (lane 2). UV-C induced marked apoptosis in cyclin I–null podocytes (lane 4). Experiments were performed in triplicate, and a minimum of 400 cells were counted per experiment. pos, positive. (B) In the absence of UV-C, Cdk5 protein levels measured by Western blot analysis were similar in nontransfected cyclin I WT and null podocytes (lanes 1, 2). siRNA reduced Cdk5 protein levels in WT cells (middle panel), which resulted in increased caspase-3 cleavage following UV-C irradiation (lower panel); caspase-3 cleavage was not increased in control siRNA–transfected cells exposed to this dose of UV-C (lanes 6, 7). β-actin served as loading control. (C) UV-C irradiation increased apoptosis (Hoechst 33342 staining) 3- to 4-fold in cyclin I WT podocytes when Cdk5 activity was inhibited by roscovitine (lane 3) compared with control cells exposed to DMSO (lane 2). (D) Roscovitine induced a progressive increase in cleaved caspase-3 (lanes 5, 6) following exposure to UV-C. β-actin served as a loading control. Data shown represent mean + SD.