A novel binding protein of single-minded 2: the mitotic arrest-deficient protein MAD2B.
Single-minded 2 (Sim2) gene, located at the Down syndrome (DS) critical region, is thought to be particularly important because of its critical role in the development of the central nervous system (CNS) and its overexpression resulting in impairment of learning and memory which is similar to that in DS. However, its exact role in DS still remains elusive. Using a yeast two-hybrid interaction trap, we identified the mitotic arrest-deficient protein MAD2B as a novel Sim2 binding protein. Through confocal laser scanning microscopy, we found that Sim2 and MAD2B colocalized in rat cortex neurons. This interaction between Sim2 and MAD2B was also confirmed by co-immunoprecipitation. Moreover, levels of mRNA and protein of both Sim2 and MAD2B were regulated during nervous development as detected by quantitative RT-PCR and western blot analysis. We also found that overexpression of Sim2 affected the distribution of MAD2B in primary culture neurons. The results of confocal laser scanning microscopy and co-immunoprecipitation indicated that in neurons, MAD2B interacted with clathrin light chain A, which is best known for its role in endocytosis. In addition, it was found that overexpression of Sim2 or MAD2B caused deficits in clathrin-mediated endocytosis. Furthermore, overexpression of both MAD2B and Sim2 exacerbates their own inhibitory effect on transferrin uptake. These results suggest that MAD2B may mediate Sim2 function during development in CNS and thereby play a critical role in pathophysiological mechanisms in DS.
Department of Neurobiology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
Xianfang
X
Meng
Xiujuan
X
Tian
Xiaolan
X
Wang
Pan
P
Gao
Chun
C
Zhang
D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics*; D001921:Brain / Q000473:pathology; D002966:Clathrin / Q000378:metabolism; D004314:Down Syndrome / Q000378:metabolism; D004705:Endocytosis; D017353:Gene Deletion; D006801:Humans; D064246:Mad2 Proteins; D018613:Microscopy, Confocal / Q000379:methods; D008938:Mitosis; D008954:Models, Biological; D009474:Neurons / Q000378:metabolism*; D011506:Proteins / Q000235:genetics*; D051381:Rats; D013997:Time Factors; D014168:Transferrin / Q000378:metabolism; D020798:Two-Hybrid System Techniques
D051792:Basic Helix-Loop-Helix Proteins; D002966:Clathrin; D064246:Mad2 Proteins; D011506:Proteins; D014168:Transferrin; C120552:MAD2L2 protein, human; C417032:SIM2 protein, human