Spatio-temporal dynamics and localization of MeCP2 and pathological mutants in living cells.
MECP2 is an X-linked gene coding for a protein functioning as a transcriptional repressor. The protein MeCP2 (Methyl CpG-binding protein) is an abundant component of pericentric heterochromatin and its mutations or duplications are present in around 80% of patients with a neurological disorder known as Rett Syndrome. Although MeCP2 action depends critically on its binding to chromatin, very little is known about the dynamics of this process. Using fluorescence recovery after photobleaching in controlled conditions concentration, we demonstrated that most GFP-MeCP2 fusion protein associates strongly and reversibly to pericentric heterocromatin whereas the remaining fraction is bound irreversibly. The mobility of the methyl-binding protein is influenced by the differentiation state of the host cells. Furthermore, residues downstream of the methyl-binding domain are critical for the interaction with chromatin. Whereas the binding is stabilised by the central region it is likely modulated by the most C-terminal region. Importantly, these residues are missing in several disease causing mutations. Our data suggest that alterations in the affinity of MeCP2 for chromatin might contribute to the pathological effects of mutations causing Rett Syndrome.
Marchi
Matilde
M
Italian Institute of Technology (IIT), Pisa, Italy.
Guarda
Alessia
A
Bergo
Anna
A
Landsberger
Nicoletta
N
Kilstrup-Nielsen
Charlotte
C
Ratto
Gian Michele
GM
Costa
Mario
M
D000818:Animals; D002454:Cell Differentiation / Q000235:genetics; D041321:Chromosomes, Human, X / Q000235:genetics / Q000378:metabolism; D006570:Heterochromatin / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D051783:Methyl-CpG-Binding Protein 2 / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D009154:Mutation; D041681:NIH 3T3 Cells; D016716:PC12 Cells; D038761:Photobleaching; D021381:Protein Transport / Q000235:genetics; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism*; D015518:Rett Syndrome / Q000235:genetics / Q000378:metabolism*
D006570:Heterochromatin; C497392:MECP2 protein, human; D051783:Methyl-CpG-Binding Protein 2; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins