The occludin and ZO-1 complex, defined by small angle X-ray scattering and NMR, has implications for modulating tight junction permeability.
Tight junctions (TJs) are dynamic cellular structures that are critical for compartmentalizing environments within tissues and regulating transport of small molecules, ions, and fluids. Phosphorylation-dependent binding of the transmembrane protein occludin to the structural organizing protein ZO-1 contributes to the regulation of barrier properties; however, the details of their interaction are controversial. Using small angle X-ray scattering (SAXS), NMR chemical shift perturbation, cross-saturation, in vitro binding, and site-directed mutagenesis experiments. we define the interface between the ZO-1 PDZ3-SH3-U5-GuK (PSG) and occludin coiled-coil (CC) domains. The interface is comprised of basic residues in PSG and an acidic region in CC. Complex formation is blocked by a peptide (REESEEYM) that corresponds to CC residues 468-475 and includes a previously uncharacterized phosphosite, with the phosphorylated version having a larger effect. Furthermore, mutation of E470 and E472 reduces cell border localization of occludin. Together, these results localize the interaction to an acidic region in CC and a predominantly basic helix V within the ZO-1 GuK domain. This model has important implications for the phosphorylation-dependent regulation of the occludin:ZO-1 complex.
Department of Biochemistry and Molecular Biology, Pennsylvania State University College of Medicine, H171, 500 University Drive, Hershey, PA 17033, USA.
Brian R
BR
Tash
Maria C
MC
Bewley
Mariano
M
Russo
Jason M
JM
Keil
Kathleen A
KA
Griffin
Jeffrey M
JM
Sundstrom
David A
DA
Antonetti
Fang
F
Tian
John M
JM
Flanagan
D000143:Acids / Q000737:chemistry; D002147:Calmodulin / Q000378:metabolism; D002463:Cell Membrane Permeability / Q000502:physiology; D004926:Escherichia coli / Q000235:genetics; D051528:Guanylate Kinases / Q000378:metabolism; D006801:Humans; D062794:MARVEL Domain Containing 2 Protein; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016296:Mutagenesis / Q000502:physiology; D019906:Nuclear Magnetic Resonance, Biomolecular; D062793:Occludin; D010750:Phosphoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation / Q000502:physiology; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D012542:Scattering, Radiation; D012996:Solutions / Q000737:chemistry; D019108:Tight Junctions / Q000378:metabolism*; D062826:Zonula Occludens-1 Protein
D000143:Acids; D002147:Calmodulin; D062794:MARVEL Domain Containing 2 Protein; C516798:MARVELD2 protein, human; D008565:Membrane Proteins; C568985:OCLN protein, human; D062793:Occludin; D010750:Phosphoproteins; D012996:Solutions; C569042:TJP1 protein, human; D062826:Zonula Occludens-1 Protein; D051528:Guanylate Kinases