Phosphorylation of tau is regulated by PKN.
For the phosphorylation state of microtubule-associated protein, tau plays a pivotal role in regulating microtubule networks in neurons. Tau promotes the assembly and stabilization of microtubules. The potential for tau to bind to microtubules is down-regulated after local phosphorylation. When we investigated the effects of PKN activation on tau phosphorylation, we found that PKN triggers disruption of the microtubule array both in vitro and in vivo and predominantly phosphorylates tau in microtubule binding domains (MBDs). PKN has a catalytic domain highly homologous to protein kinase C (PKC), a kinase that phosphorylates Ser-313 (= Ser-324, the number used in this study) in MBDs. Thus, we identified the phosphorylation sites of PKN and PKC subtypes (PKC-alpha, -betaI, -betaII, -gamma, -delta, -epsilon, -zeta, and -lambda) in MBDs. PKN phosphorylates Ser-258, Ser-320, and Ser-352, although all PKC subtypes phosphorylate Ser-258, Ser-293, Ser-324, and Ser-352. There is a PKN-specific phosphorylation site, Ser-320, in MBDs. HIA3, a novel phosphorylation-dependent antibody recognizing phosphorylated tau at Ser-320, showed immunoreactivity in Chinese hamster ovary cells expressing tau and the active form of PKN, but not in Chinese hamster ovary cells expressing tau and the inactive form of PKN. The immunoreactivity for phosphorylated tau at Ser-320 increased in the presence of a phosphatase inhibitor, FK506 treatment, which means that calcineurin (protein phosphatase 2B) may be involved in dephosphorylating tau at Ser-320 site. We also noted that PKN reduces the phosphorylation recognized by the phosphorylation-dependent antibodies AT8, AT180, and AT270 in vivo. Thus PKN serves as a regulator of microtubules by specific phosphorylation of tau, which leads to disruption of tubulin assembly.
Taniguchi
T
T
Hyogo Institute for Aging Brain and Cognitive Disorders, Himeji 670-0981, Japan. tanigu@hiabcd.go.jp
Kawamata
T
T
Mukai
H
H
Hasegawa
H
H
Isagawa
T
T
Yasuda
M
M
Hashimoto
T
T
Terashima
A
A
Nakai
M
M
Mori
H
H
Ono
Y
Y
Tanaka
C
C
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D016466:CHO Cells; D019703:Calcineurin / Q000378:metabolism; D020134:Catalytic Domain; D006224:Cricetinae; D015536:Down-Regulation; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D007166:Immunosuppressive Agents / Q000494:pharmacology; D008856:Microscopy, Fluorescence; D008870:Microtubules / Q000378:metabolism; D008969:Molecular Sequence Data; D010766:Phosphorylation; D011485:Protein Binding; D011493:Protein Kinase C / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism* / Q000502:physiology*; D017434:Protein Structure, Tertiary; D011505:Protein-Tyrosine Kinases / Q000378:metabolism* / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D012694:Serine / Q000378:metabolism; D016559:Tacrolimus / Q000494:pharmacology; D013997:Time Factors; D014162:Transfection; D014404:Tubulin / Q000378:metabolism; D014407:Tumor Cells, Cultured; D016875:tau Proteins / Q000378:metabolism*
D007166:Immunosuppressive Agents; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D014404:Tubulin; D016875:tau Proteins; D012694:Serine; D005982:Glutathione Transferase; C061570:protein kinase N; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D011493:Protein Kinase C; D019703:Calcineurin; D016559:Tacrolimus