Neuronal nuclear organization is controlled by cyclin-dependent kinase 5 phosphorylation of Ras Guanine nucleotide releasing factor-1.
RasGRF1 is a member of the Ras guanine nucleotide exchange factor (RasGEF) family of proteins which are directly responsible for the activation of Ras and Rac GTPases. Originally identified as a phosphoprotein, RasGRF1 has been shown to be phosphorylated by protein kinase A and more recently, by the non-receptor tyrosine kinases Ack1 and Src. In this report we show that RasGRF1 interacts with and is phosphorylated by Cdk5 on serine 731 to regulate its steady state levels in mammalian cells as well as in neurons. Phosphorylation on this site by Cdk5 leads to RasGRF1 degradation through a calpain-dependent mechanism. Additionally, cortical neurons from Cdk5 knockout mice have higher levels of RasGRF1 which are reduced when wild-type Cdk5 is transfected into these neurons. In mitotic cells, nuclei become disorganized when RasGRF1 is overexpressed and this is rescued when RasGRF1 is co-expressed with active Cdk5. When RasGRF1 levels are elevated in neurons through overexpression of either the wild-type RasGRF1, or the phosphorylation mutant of RasGRF1 and by the transfection of a dominant negative Cdk5 construct, nuclei appeared condensed and fragmented. On the other hand, a reduction of RasGRF1 levels through p35/Cdk5 overexpression also leads to nuclear condensation in neurons. These data show that phosphorylation of RasGRF1 by Cdk5 tightly regulates its levels, which is essential for proper cellular organization.
Kesavapany
Sashi
S
Cytoskeletal Protein Regulation Section, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD 20892, USA.
Pareek
Tej Kumar
TK
Zheng
Ya-Li
YL
Amin
Niranjana
N
Gutkind
J Silvio
JS
Ma
Wu
W
Kulkarni
Ashok B
AB
Grant
Philip
P
Pant
Harish C
HC
D000818:Animals; D001665:Binding Sites / Q000502:physiology; D016466:CHO Cells; D019556:COS Cells; D002154:Calpain / Q000378:metabolism; D016923:Cell Death / Q000235:genetics; D002467:Cell Nucleus / Q000235:genetics / Q000378:metabolism*; D002540:Cerebral Cortex / Q000378:metabolism; D002522:Chlorocebus aethiops; D006224:Cricetinae; D003412:Cricetulus; D051360:Cyclin-Dependent Kinase 5 / Q000235:genetics / Q000378:metabolism*; D015536:Down-Regulation / Q000235:genetics; D005260:Female; D005786:Gene Expression Regulation / Q000235:genetics; D051379:Mice; D018345:Mice, Knockout; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D010766:Phosphorylation; D051381:Rats; D012694:Serine / Q000378:metabolism; D015398:Signal Transduction / Q000235:genetics; D015854:Up-Regulation / Q000502:physiology; D020666:ras-GRF1 / Q000235:genetics / Q000378:metabolism*
D020666:ras-GRF1; D012694:Serine; D051360:Cyclin-Dependent Kinase 5; D002154:Calpain