Protein Kinase C (PKC)
Protein kinase C (PKC) is an AGC kinase that phosphorylates serine and threonine residues in many target proteins. It was first identified in 1977 in bovine cerebellum by Nishizuka and co-workers as a protein kinase that phosphorylated histone and protamine. Since then, its involvement in many biological processes has been demonstrated, including development, memory, differentiation, proliferation and carcinogenesis. Once thought to be a single protein, PKC is now known to comprise a large family of enzymes that differ in structure, cofactor requirements and function. Ten isoforms of PKC have been identified, varying in tissue expression and cellular compartmentalization, allowing for specific interactions with substrates.
The PKC family has been divided into three groups, differing in the enzymes' cofactor requirements; conventional (c)PKC isoforms (comprising α, βI , βII and γ), that require calcium and diacylglycerol (DAG) for activation; novel (n)PKC isoforms (comprising δ, ε, η {also known as PKC-L}, and θ that require DAG; and atypical (a)PKC isoforms, namely ζ, and ι (also known as λ, the mouse homolog of human PKCι) that require neither calcium nor DAG. Protein kinase D (PKD) is a distinct kinase family that was originally classified as a PKC subgroup (PKCμ, PKCv). The PKC-related kinases (PRK/PKN) contain kinase domains homologous to PKC's. These are not closely related to the PKC family due to very different regulatory domains; however, they can be considered to be part of the PKC superfamily.
All PKCs possess a phospholipid-binding domain for membrane interaction. The general structure of a PKC molecule consists of a catalytic and a regulatory domain, composed of a number of conserved regions, interspersed with regions of lower homology, the variable domains.
Activation of cPKCs involves translocation from the cytosol to the cell membrane by engaging the membrane-targeting modules. In the case of cPKCs, an increase in intracellular calcium first promotes the binding of the C2 domain to anionic lipids. The C1 domain binds DAG (or phorbol esters, functional DAG-analogs) and phosphatidylserine (PS), recruiting the conventional and novel PKCs to the membrane, where they can phosphorylate substrates. Specific anchoring proteins (immobilized at particular intracellular sites) localize the kinase to its site of action. These proteins include 'receptors for activated C-kinase' (RACKS), 'receptors for inactive C-kinase' (RICKS), 'A-kinase anchoring proteins' (AKAPS), and 'substrates that interact with C-kinase' (STICKS). In addition, nuclear localization signals (NLS) or nuclear export signals (NES) can send PKC into or out of the nucleus.
Some, if not all, PKC isoforms can be proteolytically cleaved at the hinge between the regulatory and catalytic domains by proteases such as the calcium-activated calpain, generating a free, cofactor-independent, catalytic subunit known as protein kinase M (PKM). This 'calpain product' should not be considered an 'unregulated' enzyme since its generation is, in fact, regulated by proteolysis. Cleavage is a physiologically relevant alternate activation mechanism for isozymes such as PKCδ, occurring in processes like apoptosis, where caspases appear to have important roles.
All PKCs, except the δ isoform, exhibit so-called PEST sequences, hydrophilic polypeptide segments enriched in proline (P), glutamic acid (E), serine (S) and threonine (T), which target proteins for degradation by the proteasome. An additional level of complexity is apparent following the observation that dephosphorylation of activated PKCs apparently predisposes them to ubiquitination and degradation. The downregulation of PKC is therefore also regulated by specific phosphatases and ubiquitin ligases.
PKC isoforms are processed by three ordered priming phosphorylations. The first phosphorylation is catalyzed by phosphoinositide-dependent kinase (PDK-1) and occurs at the activation loop (T500 in PKCβII). This phosphorylation triggers two phosphorylations at the carboxy-terminus (T641 and S660 in βII). Each phosphorylation event induces conformational changes in the PKC molecule that result in altered thermal stability, resistance to phosphatases and catalytic competency.
The first complete crystal structure of PKC was obtained for the novel isoform θ. Previous structural information was obtained from crystals of the regulatory domains, and by modeling the catalytic domain with protein kinase A.
The Table below contains accepted modulators and additional information. For a list of additional products, see the "Related Products" section below.
| Family Members/Isoforms | α (P1782) | βI (P1787) | βII (P3287) | γ (P9542) | δ (P8538) |
| Other Names | Conventional | Conventional | Conventional | Conventional | Novel |
|---|---|---|---|---|---|
| Molecular Weight (kDa) | 76.8 kDa (h) | 76.7 kDa (h) | 76.8 kDa (h) | 78.4 kDa (h) | 77.7 kDa (h) |
| Structural Data | 672 aa (h) | 671 aa (h) | 673aa (h) | 697 aa (h) | 676 aa (h) |
| Species | Ubiquitous e.g. human (h) | Ubiquitous e.g. human (h) | Ubiquitous e.g. human (h) | Ubiquitous including human | Ubiquitous including human |
| Domain Organization | N-terminal pseudosubstrate domain C1A domain C1B domain C2 domain Catalytic domain C-terminal domain | N-terminal pseudosubstrate domain C1A domain C1B domain C2 domain Catalytic domain C-terminal domain | N-terminal pseudosubstrate domain C1A domain C1B domain C2 domain Catalytic domain C-terminal domain | N-terminal pseudosubstrate domain C1A domain C1B domain C2 domain Catalytic domain C-terminal domain | N-terminal C2 domain Pseudosubstrate domain C1A domain C1B domain Catalytic domain C-terminal domain |
| Phosphorylation Sitesa | Thr497 Thr638 Ser657 (r,m,h,rb) | Thr500 Thr642 Ser661 (r,m,h,rb) | Thr500 Thr641 Ser660 (r,m,h) | Thr514 Thr655 Thr674 (r,m) | Thr505 Ser643 Ser662 (r,m,h) Thr565 Ser709 Ser728 (rb) |
| Tissue Distribution | Many; e.g. CNS, heart, kidney, liver, lung | Many; e.g. CNS, heart, kidney, liver, lung | Many; e.g. CNS, heart, kidney, liver, lung | CNS | Many; e.g. CNS, heart, kidney, liver, lung |
| Subcellular Localization | Cytosol Membrane | Cytosol Membrane | Cytosol Membrane | Cytosol Membrane | Cytosol Membrane Mitochondria |
| Binding Partners/ | All isoforms: many, e.g. RACKs, STICKs, AKAPs. See Referenceb in Footnotes. | ||||
| Associated Proteinsb | e.g. 14-3-3 | e.g. 14-3-3 Spectrin | e.g. 14-3-3 | e.g. 14-3-3 | e.g. 14-3-3 Connexin43 |
| Upstream Activators | Conventional and novel isoforms: G-protein coupled receptors, tyrosine kinase receptors, non-receptor tyrosine kinase receptors, and other agonists of phospholipid hydrolysis, atypical isoforms: binding proteins, e.g. p62/zip, par6 | ||||
| Downstream Activationc | MBP (M1891) Histones (eg. IIIS) EGFr MARCKS Protamine (P4505) AKAP79 GAP43 Pleckstrin Adducin Vinculin | MBP (M1891) Histone H3 (H4380) EGFr MARCKS Protamine (P4505) AKAP79 GAP43 Pleckstrin | MBP (M1891) Histone H3 (H4380) EGFr MARCKS Protamine (P4505) AKAP79 GAP43 Pleckstrin | MBP (M1891) Histone H3 (H4380) EGFr MARCKS Protamine (P4505) GAP43 Pleckstrin Connexin43 | MBP (M1891) Histones (eg. IIIS) EGFr Protamine (P4505) MARCKS GAP43 Pleckstrin STAT1 STAT3 Adducin |
| Substrates | α Pseudosubstrate peptide MBP peptide (P2186) Syntide 2 EGFr peptide (L9905) PKC substrate MARCKS peptide | α Pseudosubstrate peptide MBP peptide (P2186) Syntide 2 EGFr peptide (L9905) PKC substrate MARCKS peptide | α Pseudosubstrate peptide MBP peptide (P2186) Syntide 2 EGFr peptide (L9905) PKC substrate MARCKS peptide | α Pseudosubstrate peptide MBP peptide (P2186) Syntide 2 EGFr peptide (L9905) PKC substrate MARCKS peptide | α Pseudosubstrate peptide ε Pseudosubstrate peptide MBP peptide (P2186) EGFr peptide (L9905) PKC substrate eEF-1α MARCKS peptide |
| Activatorsd | Ca2+ DAG (O6754) PDA (P9143) PDBu (P1269) PDD (P9018) PMA (P8139) PS (P5660, P6641, P7769) | Ca2+ DAG (O6754) PDA (P9143) PDBu (P1269) PDD (P9018) PMA (P8139) PS (P5660, P6641, P7769) | Ca2+ DAG (O6754) PDA (P9143) PDBu (P1269) PDD (P9018) PMA (P8139) PS (P5660, P6641, P7769) | Ca2+ DAG (O6754) PDA (P9143) PDBu (P1269) PDD (P9018) PMA (P8139) PS (P5660, P6641, P7769) | DAG (O6754) PDA (P9143) PDBu (P1269) PDD (P9018) PMA (P8139) PS (P5660, P6641, P7769) |
| Inhibitorse | Calphostin C (C6303) Ro 31-8220 (R136) GF109203X (G2911) Gö 6976 (G1171) K252a (K1639, K2015) Ro 31-7549 Gö 6983 (G1918) Chelerythrine chloride (C2932) (–)-Balanol UCN-01 (U6508) CGP41251 CGP54345 CGP53506 Aprinocarsen CGP53506 | Calphostin C (C6303) Ro 31-8220 (R136) GF109203X (G2911) Gö 6976 (G1171) K252a (K1639, K2015) Ro 31-7549 Gö 6983 (G1918) Chelerythrine chloride (C2932) (–)-Balanol UCN-01 (U6508) CGP41251 LY-333531 (SML0693) LY379196 LY317615 (SML0762) Aprinocarsen | Calphostin C (C6303) Ro 31-8220 (R136) GF109203X (G2911) Gö 6976 (G1171) K252a (K1639, K2015) Ro 31-7549 Gö 6983 (G1918) (–)-Balanol UCN-01 (U6508) CGP41251 LY-333531 (SML0693) LY379196 LY317615 (SML0762) | Calphostin C (C6303) Ro 31-8220 (R136) GF109203X (G2911) Gö 6976 (G1171) K252a (K1639, K2015) Ro 31-7549 Gö 6983 (G1918) Chelerythrine chloride (C2932) (–)-Balanol UCN-01 (U6508) CGP41251 | Calphostin C (C6303) Gö 6983 (G1918) Chelerythrine chloride (C2932) LY-333351 (SML0693) (–)-Balanol Rottlerin (R5648) |
| Selective Activators | Not Known | Not Known | Not Known | Not Known | Not Known |
| Physiological Function | Cell motility Migration Apoptosis | Cell growth Immune function B-cell survival | Cell growth Immune function B-cell survival Antiapoptotic | Nociceptive response Spatial learning Neuroplasticity | Proapoptotic Growth Differentiation Platelet granular secretion |
| Disease Relevance | Cardiac hypertrophy Heart failure Prostate cancer Breast cancer Chemo therapeutic resistance | Cardiac hypertrophy Heart failure Gastric cancer | Cardiac hypertrophy Heart failure Leukemia Colon cancer | Spinocerebellar ataxia Lymphoma | Cardiac hypertrophy Cardiac ischemic injury Prostate cancer Leukemia |
| Family Members/Isoforms | ε | η/L | θ | ζ | ι/λ |
|---|---|---|---|---|---|
| Other Names | Novel | Novel | Novel | Atypical | Atypical |
| Molecular Weight (kDa) | 83.5 kDa (h) | 77.9 kDa (h) | 81.9 kDa (h) | 67.7 kDa (h) | 67.3 kDa (h) |
| Structural Data | 737 aa (h) | 680 aa (h) | 706 aa (h) | 592 aa (h) | 587 aa (h) |
| Species | Ubiquitous including human | Ubiquitous e.g. human | Ubiquitous e.g. human | Ubiquitous e.g. human | Ubiquitous e.g. human |
| Domain Organization | N-terminal C2 domain Pseudosubstrate domain C1A domain C1B domain Catalytic domain C-terminal domain | N-terminal C2 domain Pseudosubstrate domain C1A domain C1B domain Catalytic domain C-terminal domain | N-terminal C2 domain Pseudosubstrate domain C1A domain C1B domain Catalytic domain C-terminal domain | N-terminal PB1 domain Pseudosubstrate domain Atypical C1 domain Catalytic domain C-terminal domain | N-terminal domain PB1 domain Pseudosubstrate domain Atypical C1 domain Catalytic domain C-terminal domain |
| Phosphorylation Sitesa | Thr566 Thr710 Ser729 (h,r) | Thr512 Thr655 Ser674 (h) Thr513 Thr656 Ser675 (r,m) | Thr538 Ser676 Ser695 (m,h) | Thr410 Thr560 (r,m,h) | Thr412 Thr564 (h) |
| Tissue Distribution | Many; e.g. CNS, heart, kidney, liver, lung | Many; e.g. CNS, heart, lung, spleen | Many; e.g. CNS, heart, liver, airway smooth muscle, lung | Many; e.g. CNS, heart, kidney, liver, lung | Many; e.g. CNS, heart, airway smooth muscle, liver, lung |
| Subcellular Localization | Cytosol Membrane Golgi | Cytosol Membrane | Cytosol Membrane | Cytosol Membrane | Cytosol Membrane |
| Binding Partners/ | All isoforms: many, e.g. RACKs, STICKs, AKAPs. See Referenceb in Footnotes. | ||||
| Associated Proteinsb | e.g. integrins Connexin43 | e.g. cdk2 | e.g. IKKβ | e.g. p62/ZIP Par6 mek5 | e.g. p62 ZIP5 mek5 LIP Par6 |
| Upstream Activators | Conventional and novel isoforms: G-protein coupled receptors, tyrosine kinase receptors, non-receptor tyrosine kinase receptors, and other agonists of phospholipid hydrolysis, atypical isoforms: binding proteins, e.g. p62/zip, par6 | ||||
| Downstream Activationc | MBP (M1891) Histones EGFr Protamine (P4505) MARCKS GAP43 Pleckstrin | MBP (M1891) Histones EGFr Protamine (P4505) MARCKS GAP43 Pleckstrin Integrin βII | MBP (M1891) Histones EGFr Protamine (P4505) MARCKS GAP43 Pleckstrin | MBP (M1891) Histones EGFr Protamine (P4505) Pleckstrin | MBP (M1891) Histones EGFr Protamine (P4505) Pleckstrin MARCKS |
| Substrates | α Pseudosubstrate peptide ε Pseudosubstrate peptide (P4459) ε Substrate MBP peptide (P2186) EGFr peptide (L9905) PKC substrate MARCKS peptide | η Substrate α Pseudosubstrate peptide ε Pseudosubstrate peptide (P4459) MBP peptide (P2186) EGFr peptide (L9905) PKC substrate MARCKS peptide | θ Substrate α Pseudosubstrate peptide MBP peptide (P2186) EGFr peptide (L9905) PKC substrate MARCKS peptide | ε Substrate ε Pseudosubstrate peptide (P4459) α Pseudosubstrate peptide MBP peptide (P2186) EGFr peptide (L9905) PKC substrate | ε Pseudosubstrate peptide (P4459) α Pseudosubstrate peptide MBP peptide (P2186, M8184) EGFr peptide (L9905) PKC substrate MARCKS peptide |
| Activatorsd | DAG (O6754) PDA (P9143) PDBu (P1269) PDD (P9018) PMA (P8139) PS (P5660, P6641, P7769) | DAG (O6754) PDA (P9143) PDBu (P1269) PDD (P9018) PMA (P8139) PS (P5660, P6641, P7769) | DAG (O6754) PDA (P9143) PDBu (P1269) PDD (P9018) PMA (P8139) PS (P5660, P6641, P7769) | PS (P5660, P6641, P7769) p62/ZIP Par6 | PS (P5660, P6641, P7769) p62/ZIP Par6 |
| Inhibitorse | Calphostin C (C6303) GF109203X (G2911) Gö 6983 (G1918) LY-333531 (SML0693) (–)-Balanol | Calphostin C (C6303) GF109203X (G2911) Gö 6983 (G1918) LY-333351 (SML0693) (–)-Balanol η pseudosubstrate peptide NPC 15437 (N161) | Calphostin C (C6303) GF109203X (G2911) Gö 6983 (G1918) LY-333531 (SML0693) (–)-Balanol θ pseudosubstrate peptide | Gö 6983 (G1918) ζ pseudosubstrate peptide (P1614) | Gö 6983 (G1918) |
| Selective Activators | Not Known | Not Known | Not Known | Not Known | Not Known |
| Physiological Function | Cell migration Antiapoptotic Nociceptive response Macrophage activation | Antiapoptotic Cell cycle Differentiation B-cell development | Immune function (T-cell signaling) Cytoskeletal assembly Antiapoptotic | Immune function (B-cell signaling) Cell polarity Antiapoptotic | Cell polarity Antiapoptotic Insulin sensitivity |
| Disease Relevance | Cardiomyopathy Ischemic preconditioning Alzheimer’s disease Lung cancer Colorectal cancer Skin cancer Diabetes | Glioblastoma Lung cancer Renal cancer Skin cancer Breast cancer | Gastrointestinal cancer Breast cancer Leukemia | Colon cancer Pancreatic cancer Renal cancer Leukemia | Colon cancer Leukemia Diabetes |
Footnotes
a) Processing sites listed.
b) For detailed list of binding partners, see Reference: Poole, A.W., et al. “PKC-interacting proteins: from function to pharmacology.” Trends Pharmacol Sci, 25, 528-535 (2004).
c) PKCα pseudosubstrate site sequence RFARKGALRQKNVHEVKDH. PKCε pseudosubstrate site sequence PRKRQGAVRRRVHQVNGH (underlined are mutable sites for phosphorylateable residues). Substrates not determined on all isoforms.
d) Specific lipid activator of aPKCs unknown. Less is known about activators/substrates/inhibitors of aPKCs than nPKCs than cPKCs. Activators not determined on all isoforms.
e) GF 109203X (Gö 6850 aka BIM 1) displays potency rank order of α>βI>ε>δ>ζ. Bisindolylmaleimides (Ro compounds, BIM 1, Gö 6976) inhibit all PKCs in potency rank order of cPKCs>nPKCs>aPKCs (generally); Calphostin C inhibits DAG site in regulatory domain; BIM 1 and Ro compounds act at ATP site; Chelerythrine chloride inhibits substrate site in catalytic domain.
Abbreviations
AKAP: A-Kinase Anchoring Protein
Aprinocarsen: ISIS 3521
Cdk2: Cyclin dependent kinase 2
CGP41251: (PKC412): Staurosporine derivative
CGP54345: ATP analog
CGP53506: N-(3-Nitrophenyl)-4-(3-pyridyl)-2-pyrimidinamine
eEF-1α: Eukaryotic elongation factor-1α peptide, residues 422-443.
Gö 6976: 5,6,7,13-Tetrahydro-13-methyl-5-oxo-12H-indolo[2,3-a]pyrrolo[3,4-c]carbazole-12-propanenitrile
Gö 6983: 2-[1-(3-Dimethylaminopropyl)-5-methoxyindol-3-yl]-3-(1H-indol-3-yl)maleimide
GF 109203X: 3-[1-[3-(Dimethylamino)propyl]-1H-indol-3-yl]-4-(1H-indol-3-yl)-1H-pyrrole-2,5-dione (Gö6850)
LIP: PKC 1 interacting protein to PKC λ interacting protein
LY-333531: 9-[(Dimethylamino)methyl]-6,7,10,11-tetrahydro-(9S)-NH,18H-5,21:12,15-dimetheno-dibenzo[e,k]pyrrolo[3,4-h][1,4,13]oxadiazacyclohexadecine-18,20(19H)dione
K252a: Staurosporine-related alkaloid
MARCKS peptide: Myristoylated alanine-rich C-kinase substrate, Ac-phe-lys-lys-ser-phe-lys-leu-NH2
MBP: Myelin basic protein
NPC 15437: S-2,6-Diamino-N-[[1'-(1''oxotridecyl)-2'-piperidinyl]methyl]hexanamide dihydrochloride
PB1: Phox and Bem1p binding domain
PDA: Phorbol 12, 13-diacetate
PDBu: Phorbol 12, 13-dibutyrate
PDD: Phorbol 12, 13-didecanoate
Pleckstrin: Platelet and leukocyte C-kinase substrate protein
PMA: Phorbol 12-myristate 13-acetate
PS: Phosphatidylserine
RACK: Receptors for Activated C-Kinase
Ro 31-7549: 2-[1-3(Aminopropyl)indol-3-yl]-3(1-methyl-1H-indol-3-yl)maleimide
Ro 31-8220: 2-{1-[3-(Amidinothio)propyl]-1H-indol-3-yl}-3-(1-methylindol-3-yl)-maleimide)
STAT: Signal Transducers and Activators of Transcription
STICK: Substrates That Interact with C-Kinase
Syntide 2: H-pro-leu-ala-arg-thr-leu-ser-val-ala-gly-leu-pro-gly-lys-lys-OH
TPA: Tetra phorbol acetate
UCN-01: 7-Hydroxy-staurosporine
ZIP: PKC Zeta interacting protein
r: Rat
h: Human
m: Mouse
rb: Rabbit
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References
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