The Rho GTPase and Fyn tyrosine kinase have been implicated previously in positive control of keratinocyte cell–cell adhesion. Here, we show that Rho and Fyn operate along the same signaling pathway. Endogenous Rho activity increases in differentiating keratinocytes and is required for both Fyn kinase activation and increased tyrosine phosphorylation of β- and γ-catenin, which is associated with the establishment of keratinocyte cell–cell adhesion. Conversely, expression of constitutive active Rho is sufficient to promote cell–cell adhesion through a tyrosine kinase- and Fyn-dependent mechanism, trigger Fyn kinase activation, and induce tyrosine phosphorylation of β- and γ-catenin and p120ctn. The positive effects of activated Rho on cell–cell adhesion are not induced by an activated Rho mutant with defective binding to the serine/threonine PRK2/PKN kinases. Endogenous PRK2 kinase activity increases with keratinocyte differentiation, and, like activated Rho, increased PRK2 activity promotes keratinocyte cell–cell adhesion and induces tyrosine phosphorylation of β- and γ-catenin and Fyn kinase activation. Thus, these findings reveal a novel role of Fyn as a downstream mediator of Rho in control of keratinocyte cell–cell adhesion and implicate the PRK2 kinase, a direct Rho effector, as a link between Rho and Fyn activation.
Nodular pulmonary amyloidosis, a subtype of pulmonary amyloidosis, is a unique disease that can mimic lung cancer on radiographic imaging and is related to lymphoproliferative disorders. In this report, we describe a case of a 76-year-old male who presented with a solitary nodule in his left lower lung lobe on computed tomography that increased from 6 mm to 13 mm in diameter over 40 months. Lung cancer was suspected; however, transbronchial lung biopsy revealed deposition of an eosinophilic and homogeneous amorphous substance, which showed apple-green birefringence under polarized light after Congo red staining, and immunohistochemistry analysis returned positive results for immunoglobulin lambda light-chain. Upper gastrointestinal endoscopy revealed a gastric mucosa-associated lymphoid tissue (MALT) lymphoma. These findings indicated that this was a case of nodular pulmonary amyloidosis that preceded a diagnosis of MALT lymphoma.
PKN is a fatty acid- and Rho GTPase-activated protein kinase whose catalytic domain in the carboxyl terminus is homologous to those of protein kinase C (PKC) family members. The amino terminal region of PKN is suggested to function as a regulatory domain, since tryptic cleavage or the binding of Rho GTPase to this region results in protein kinase activation of PKN. The structural basis for the regulation of PKN was investigated by analyzing the activity of a series of deletion/site-directed mutants expressed in insect cells. The amino-terminally truncated form of PKN (residue 455–942) showed low basal activity similar to that of the wild-type enzyme, and was arachidonic acid-dependent. However, further deletion (residue 511–942) resulted in a marked increase in the basal activity and a decrease in the arachidonic acid dependency. A (His)6 -tagged protein comprising residues 455–511 of PKN (designated His-Ia) inhibited the kinase activity of the catalytic fragment of PKN in a concentration-dependent manner in competition with substrate (K1=0.6±0.2 μM). His-Iα also inhibited the activity of the catalytic fragment of PRK2, an isoform of PKN, but had no inhibitory effect on protein kinase A or protein kinase C8. The IC50 value obtained in the presence of 40 μM arachidonic acid was two orders of magnitude greater than that in the absence of the modifier. These results indicate that this protein fragment functions as a specific inhibitor of PKN and PRK2, and that arachidonic acid relieves the catalytic activity of wild-type PKN from autoinhibition by residues 455–511 of PKN. Autophosphorylation of wild-type PKN increased the protein kinase activity, however, substitution of Thr64, Ser374, or Thr531 in the regulatory region of PKN with alanine, abolished this effect. Substitution of Thr774 in the activation loop of the catalytic domain of PKN with alanine completely abolished the protein kinase activity. These results suggest that these phosphorylation sites are also important in the regulation of the PKN kinase activity. Potential differences in the mechanism of activation between the catalytic regions of PKN and PRK2 are also discussed.
