Identification of ZASP, a novel protein associated to Zona occludens-2.
With the aim of discovering new molecular interactions of the tight junction protein ZO-2, a two-hybrid screen was performed on a human kidney cDNA library using as bait the middle segment of ZO-2. Through this assay we identified a 24-kDa novel protein herein named ZASP for ZO-2 associated speckle protein. ZO-2/ZASP interaction further confirmed by pull down and immunoprecipitation experiments, requires the presence of the intact PDZ binding motif SQV of ZASP and the third PDZ domain of ZO-2. ZASP mRNA and protein are present in the kidney and in several epithelial cell lines. Endogenous ZASP is expressed primarily in nuclear speckles in co-localization with splicing factor SC-35. Nocodazole treatment and wash out reveals that ZASP disappears from the nucleus during mitosis in accordance with speckle disassembly during metaphase. ZASP amino acid sequence exhibits a canonical nuclear exportation signal and in agreement the protein exits the nucleus through a process mediated by exportin/CRM1. ZASP over-expression blocks the inhibitory activity of ZO-2 on cyclin D1 gene transcription and protein expression. The identification of ZASP helps to unfold the complex nuclear molecular arrays that form on ZO-2 scaffolds.
Lechuga
Susana
S
Department of Physiology, Biophysics and Neuroscience, Center for Research and Advanced Studies (Cinvestav), México, DF 07360, Mexico.
Alarcón
Lourdes
L
Solano
Jesús
J
Huerta
Miriam
M
Lopez-Bayghen
Esther
E
González-Mariscal
Lorenza
L
D048868:Adaptor Proteins, Signal Transducing / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D018949:Antigens, CD1 / Q000378:metabolism; D001483:Base Sequence; D002453:Cell Cycle; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D003001:Cloning, Molecular; D019938:Cyclin D1 / Q000235:genetics; D004285:Dogs; D005786:Gene Expression Regulation; D006801:Humans; D047468:Immunoprecipitation; D028884:Karyopherins / Q000378:metabolism; D007668:Kidney / Q000378:metabolism; D060588:LIM Domain Proteins; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D062827:Zonula Occludens-2 Protein
D048868:Adaptor Proteins, Signal Transducing; D018949:Antigens, CD1; D028884:Karyopherins; C120696:LDB3 protein, human; D060588:LIM Domain Proteins; D008565:Membrane Proteins; D012333:RNA, Messenger; C569046:TJP2 protein, human; D062827:Zonula Occludens-2 Protein; D019938:Cyclin D1