Tuberous sclerosis complex (TSC) is a tumor suppressor gene syndrome caused by 
mutations in TSC1 and TSC2. Hamartin and tuberin, the products of TSC1 and TSC2, 
respectively, form heterodimers and inhibit the mammalian target of rapamycin. 
Previously, we have shown that hamartin is phosphorylated by CDC2/cyclin B1 
during the G(2)/M phase of the cell cycle. Here, we report that hamartin is 
localized to the centrosome and that phosphorylated hamartin and phosphorylated 
tuberin co-immunoprecipitate with the mitotic kinase Plk1. Plk1 interacts with 
the N-terminus of hamartin (amino acids 1-880), which contains two potential 
Plk1-binding sites (T310 and S332). Phosphorylated hamartin interacts with Plk1 
independent of tuberin with all three proteins present in a complex. A 
non-phosphorylatable hamartin mutant with an alanine substitution at residue 
T310 does not interact with Plk1, whereas a non-phosphorylatable hamartin mutant 
at residue S332 in conjunction with alanine mutations at the other CDC2/cyclin 
B1 sites (T417, S584 and T1047) does not impact hamartin binding to Plk1. 
Hamartin negatively regulates the protein levels of Plk1. Finally, Tsc1(-/-) 
mouse embryonic fibroblasts (MEFs) have increased number of centrosomes and 
increased DNA content, compared to Tsc1(+/+) cells. Both phenotypes are rescued 
after pre-treatment with the mTOR inhibitor rapamycin. RNAi inhibition of Plk1 
in Tsc1(-/-) MEFs failed to rescue the increased centrosome number phenotype. 
These data reveal a novel subcellular localization for hamartin and a novel 
interaction partner for the hamartin/tuberin complex and implicate hamartin and 
mTOR in the regulation of centrosome duplication.