Other kinases

Many other kinases and cofactors have been shown to undergo important translocations in nonmuscle cells, but in general these observations have not yet been extended to differentiated smooth muscle. 

Raf-1 is a serine/threonine kinase that functions by phosphorylating and activating yet other kinases such as MAP kinase kinase (Kyriakis et al., 1992). Interestingly, Raf-1 may also function as a substrate for PKC (Macdonald et al., 1993 Kokh et al., 1993), a process that may control the growth of undifferentiated cells. The fact that Raf-1 is also abundant in differentiated cell types, particularly smooth muscle (Adam and Hathaway, 1993), and that it can be activated by PKC (Kokh etal., 1993) suggest that Raf-1 may play a signaling role in contractile smooth muscle cells. 

In unstimulated macrophages, 90% of the total activity of phospholipase A2 is localized to the cytosol. l-Oleoyl-2-acetyl-glycerol (OAG) causes translocation of phospholipase A2 to the membrane fraction (Schon-hardt and Ferber, 1987). Also, an increase in Ca + concentration has been shown to induce translocation of phospholipase A2 from the cytosolic fraction to the membrane fraction of rat brain (Yoshihara and Wat-anabe, 1990) and rat liver (Krause et al., 1991). 

Evidence in the literature suggests that MLC kinase (MLCK) is permanently targeted to the vicinity of the light chains (Adelstein and Klee, 1981 Sobieszek, 

and does not undergo translocations. However, this is largely based on the fact that the kinase is found bound to the insoluble fraction during purification procedures. To the best of our knowledge, studies directly assessing the subcellular location of MLCK in situ have not been performed. 

In addition to these kinases, there are undoubtedly others in neutrophils, including Ca2+/calmodulin-activated kinases, which regulate function. Furthermore, there are a variety of phosphatases that can reverse the effects 

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In most animal, plant, and microbial cells, the enzyme that phosphorylates glucose is hexokinase. Magnesium ion (Mg ) is required for this reaction, as for the other kinase enzymes in the glycolytic pathway. The true substrate for the hexokinase reaction is MgATP. The apparent K , for glucose of the animal... 

Several other inhibitors of nonreceptor PTKs are currently in development but only a few of them are studied in clinical trials. Noteworthy, Dasatinib does not only inhibit c-Abl, but also potently blocks Src activity, a property that may contribute to its beneficial clinical effects in CML. Other kinase inhibitors being developed that inhibit c-Abl and/or Src are AZD-0530, AP-23994, PD-0183805, SU-6656, and Bosutinib (SKI-606). Furthermore, peptidomimetic SH2 domain inhibitors for Src, such as AP-22408 have been designed that decrease bone resorption and may be promising drugs to treat osteoporosis and other bone diseases, such as Paget s disease and osteolytic bone metastasis. 

To date, no specific inhibitors ofMK2 have been reported. However, since MKT is activated by p38 MAP kinases a// , use of SB203580 will prevent its activation (but also interferes with the control of other kinases, such as Mnkla, that are activated by the same pathway). 

C-termini and a large glycosylated extracellular loop between transmembrane domains 3 and 4. The proteins show the most homology in their transmembrane spanning domains, particularly domains 1, 2, and 4-8, which may be involved in moving the transmitter across the membrane. The transporters are substrates for PKC-dependent phosphorylation, which reduces their activity. The dopamine transporter is phosphorylated on the extreme end of the N-terminal tail, and consensus phosphorylation sites for various other kinases are present in the intracellular loops and domains [20-22] (Fig. 12-4). The dopamine and norepinephrine transporters form functional homo-oligomers, although it is not known if this is required for transport activity, and the transporters also interact with many other membrane proteins that may control their cell-surface expression or other properties. 

ARRY-438162 is a recently disclosed potent and selective ATP non-competitive MEK1/2 inhibitor that is in Phase lb clinical trials as an anti-arthritic agent [24]. ARRY-438162 inhibited the MEK1/2 enzyme with an IC50 = 12 nM and pERK in cells with an IC50 = 11 nM. ATP non-competitive inhibition may be responsible for equipotent inhibition of MEK1/2 in vitro and pERK in cells. The compound was selective against a panel of 220 other kinases. 

AS601245 has been reported to be an ATP-competitive inhibitor of JNK1, 2 and 3 with IC50 values = 150, 220 and 70 nM, respectively, with minimal to no activity in a panel of 25 other kinases [48]. In Jurkat T-cells, AS601245 at 10 pM inhibited IL-2 production induced by phorbol-12-myristate-l 3-acetate (PMA) by 90%. The weaker cell activity could be due to poor cell permeability. The oral bioavailability of AS601245 in rats was 38%. In mice, AS601245, when dosed at 60 mg/kg p.o. in a developed arthritis model induced by collagen, showed... 

Other kinase receptors are serine/threonine kinases, protein kinases, and mitogen-activated protein (MAP) kinases. Insulin, transforming growth factor-beta (TGF- 8), and platelet-derived growth factor (PDGF) are the natural ligands that interact with kinase receptors. 

The invariant SQ motif in the C-terminus of H2AX is a consensus sequence for the 3 kinases belonging to the PIKK family, namely ATM, DNA-PK and ATR (Stiff et al, 2004). These kinases are involved in DNA repair. ATM [ataxia telangiectasia (A-T) mutated protein] is a crucial kinase for the signal transduction DSB pathway (Savitsky et al, 1995) and it is widely accepted that ATM is the major kinase involved in the in vivo phosphorylation of H2AX (Burma et al, 2001 Fernandez-Capetillo et al, 2002 Redon et al, 2002). The two other kinases were also associated with the generation y-H2AX, but they appeared not to be dominant (Redon et al, 2002 Stiff et al, 2004). 

AG 1024 has been extensively studied as an IGFR inhibitor [70] and is a substrate competitive inhibitor of this kinase [71]. AG1024 also inhibits other kinases including c-Kit [72]. Additional studies will be needed, including a direct measurement of Abl activity and possible subsequent testing against the imatinib resistant Abl point mutations, to ascertain the possible therapeutic utility of AG 1024. 

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