Members of the mammalian protein kinase C (PKC) superfamily play key regulatory roles in a multitude of cellular processes, ranging from control of fundamental cell autonomous activities, such as proliferation, to more organismal functions, such as memory. However, understanding of mammalian PKC signalling systems is complicated by the large number of family members. Significant progress has been made through studies based on comparative analysis, which have defined a number of regulatory elements in PKCs which confer specific location and activation signals to each isotype. Further studies on simple organisms have shown that PKC signalling paradigms are conserved through evolution from yeast to humans, underscoring the importance of this family in cellular signalling and giving novel insights into PKC function in complex mammalian systems.

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The extended protein kinase C superfamily.

Ribosome-inactivating proteins from plants

https://febs.onlinelibrary.wiley.com/doi/epdf/10.1016/0014-5793%2896%2900785-5

Inhibition of protein kinase C μ by various inhibitors. Inhibition from protein kinase c isoenzymes

Protein kinase C (PKC), a family of related serine-threonine kinases, is a key player in the cellular responses mediated by the second messenger diacylglycerol (DAG) and the phorbol ester tumor promoters. The traditional view of PKCs as DAG/phospholipid-regulated proteins has expanded in the last few years by three seminal discoveries. First, PKC activity and maturation is controlled by autophosphorylation and transphosphorylation mechanisms, which includes phosphorylation of PKC isozymes by phosphoinositide-dependent protein kinases (PDKs) and tyrosine kinases. Second, PKC activity and localization are regulated by direct interaction with different types of interacting proteins. Protein-protein interactions are now recognized as important mechanisms that target individual PKCs to different intracellular compartments and confer selectivity by associating individual isozymes with specific substrates. Last, the discovery of novel phorbol ester receptors lacking kinase activity allows us to speculate that some of the biological responses elicited by phorbol esters or by activation of receptors coupled to elevation in DAG levels could be mediated by PKC-independent pathways.

New insights into the regulation of protein kinase C and novel phorbol ester receptors

Protein kinase C (PKC) isozymes comprise a family of related enzymes. There are only limited differences between these isozymes in substrate specificity or sensitivity to activators. However, there are multiple isozymes within a cell mediating isozyme-specific functions. Differential subcellular localization has been proposed to explain this specificity. When members of the PKC family are activated by lipid-derived second messengers, they translocate from one cell compartment to another. Isozyme specificity appears to be mediated in part by association of each PKC isozyme with specific anchoring proteins. This review will cover the proteins involved in the anchoring of PKC isozymes at specific subcellular sites, the domains in the PKC isozymes that mediate protein-protein interaction with isozyme-specific anchoring proteins, and identification of peptides that interfere with or promote these protein-protein interactions, thus altering the localization and function of individual isozymes.

https://faseb.onlinelibrary.wiley.com/doi/pdf/10.1096/fsb2fasebj.12.1.35

Anchoring proteins for protein kinase C: a means for isozyme selectivity

PKCu is a novel, atypical member of the protein kinase C family.

Protein kinase C mu (PKC mu) displays unusual structural features like a pleckstrin homology domain and an amino-terminal hydrophobic region with a putative leader peptide and transmembrane sequence. As a discrete location often is a direct clue to the potential biological function of a kinase, antibodies directed against unique amino- and carboxy-terminal domains of PKC mu were used to localize the protein within intracellular compartments in immunofluorescence and subcellular fractionation studies. Confocal laser scanning microscopy showed colocalization of PKC mu with the resident Golgi marker protein beta 1,4 galactosyltransferase in PKC mu transfectants and in the human hepatocellular carcinoma cell line HepG2, expressing endogenous PKC mu. Long-term treatment of cells with brefeldin A, which disintegrates the Golgi apparatus, disrupted PKC mu-specific staining. Cosegregation of PKC mu with beta 1,4 galactosyltransferase, but not with the endosomal marker rab5, upon density gradient fractionation and Western blot analysis of HepG2 cell extracts, provides independent evidence for a Golgi localization of PKC mu. Moreover, cellular sulfate uptake and Golgi-specific glycosaminoglycan sulfation was enhanced in PKC mu transfectants. Together, these data suggest that PKC mu is a resident protein kinase of the core Golgi compartment and is involved in basal transport processes.

https://rupress.org/jcb/article-pdf/134/6/1401/1256437/1401.pdf

Protein kinase C mu is located at the Golgi compartment.

In order to investigate regulatory mechanisms and to identify potential substrates of a novel member of the protein kinase C (PKC) family, PKC mu, specific antibodies have been raised against unique amino- and carboxy-terminal regions. PKC mu kinase activity was studied upon immunoprecipitation from stably transfected cell lines as well as from the A549 carcinoma cell line expressing the endogenous PKC mu gene. Cell fractionation revealed that PKC mu is predominantly found in the particulate fraction, suggesting an association with the membrane or membrane-bound structures. In vitro kinase assays with immunoprecipitated PKC mu demonstrated a Ca2+ independent enhancement of constitutive autophosphorylation activity by phosphatidylserine. Despite a limited in vitro phorbol ester response, an apparent phorbol ester activation of PKC mu was observed when cell cultures, instead of immunoprecipitated enzyme, were treated with either phorbol 12-myristate 13-acetate or 1,2 dioleoyl-sn-glycerol. Both in vitro autophosphorylation and substrate phosphorylation of myelin basic protein and histone III were enhanced under these conditions. However, long-term treatment with the phorbol ester did not result in downregulation of PKC mu protein levels and kinase activity. Studies with several protein kinase inhibitors revealed a novel sensitivity profile of PKC mu, with no inhibition by calphostin C, reduced sensitivity to staurosporine but, compared to other PKCs, an approximately 60-fold higher sensitivity to the selective PKA inhibitor H89. Together, the data presented here show that localization of PKC mu and regulation of its kinase activity differ from that of other PKCs suggesting a novel function of PKC mu in intracellular signal pathways.

https://febs.onlinelibrary.wiley.com/doi/pdf/10.1111/j.1432-1033.1995.tb20389.x

Characterization of Activators and Inhibitors of Protein Kinase Cμ

Four different type 1 ribosome-inactivating proteins (RIPs) with RNA N-glycosidase activity were tested for their ability to attack the large rRNA of plant ribosomes derived from tobacco plants, as well as from the plant species from which the particular RIP had been isolated. Incubation of tobacco ribosomes with RIPs isolated from either Phytolacca americana L. (pokeweed), Dianthus barbatus L. (carnation), Spinacia oleracea L. (spinach) or Chenopodium amaranthicolor Coste and Reyn. (chenopodium) rendered the 25S rRNA sensitive to aniline-catalyzed hydrolysis, generating a single rRNA-fragment of about 350 nucleotides. The same fragment was generated when rRNAs from pokeweed, carnation, spinach or chenopodium ribosomes were aniline-treated without any deliberate treatment of the ribosomes with the respective RIP. This indicated that ribosomes from all RIP-producing plants were already inactivated by their own RIPs during preparation. These results demonstrate that plant ribosomes are generally susceptible to RIP attack, including modification by their own RIPs. Direct sequencing of the newly generated fragments revealed that a single N-glycosidic bond at an adenosine residue within the highly conserved sequence 5'-AGUACGAGAGGA-3' was cleaved by all of the RIPs investigated, a situation also found in animal, yeast and Escherichia coli ribosomes.

J. Prestle

Papers

PKCu is a novel, atypical member of the protein kinase C family.

Protein kinase C mu is located at the Golgi compartment.

Characterization of Activators and Inhibitors of Protein Kinase Cμ

Type 1 ribosome-inactivating proteins depurinate plant 25S rRNA without species specificity

Mechanism and site of action of a ribosome-inactivating protein type 1 from Dianthus barbatus which inactivates Escherichia coli ribosomes.