Article

Cyclic AMP-Independent Regulation of Protein Kinase A Substrate Phosphorylation by Kelch Repeat Proteins

Department of Pharmacology and Biological Chemistry, Mount Sinai School of Medicine, Box 1603, 1 Gustave L. Levy Place, New York, NY 10029, USA.
Eukaryotic Cell (Impact Factor: 3.18). 12/2005; 4(11):1794-800. DOI: 10.1128/EC.4.11.1794-1800.2005
Source: PubMed
ABSTRACT
Pseudohyphal and invasive growth in the yeast Saccharomyces cerevisiae is regulated by the kelch repeat-containing proteins Gpb1p and Gpb2p, which act downstream of the G protein α-subunit Gpa2p.
Here we show that deletion of GPB1 and GPB2 causes increased haploid invasive growth in cells containing any one of the three protein kinase A (PKA) catalytic subunits,
suggesting that Gpb1p and Gpb2p are able to inhibit each of these kinases. Cells containing gpb1Δ gpb2Δ mutations also display increased phosphorylation of the PKA substrates Sfl1p and Msn2p, indicating that Gpb1p and Gpb2p
are negative regulators of PKA substrate phosphorylation. Stimulation of PKA-dependent signaling by gpb1Δ gpb2Δ mutations occurs in cells that lack both adenylyl cyclase and the high-affinity cyclic AMP (cAMP) phosphodiesterase. This
effect is also seen in cells that lack the low-affinity cAMP phosphodiesterase. Given that these three enzymes control the
synthesis and degradation of cAMP, these results indicate that the effect of Gpb1p and Gpb2p on PKA substrate phosphorylation
does not occur by regulating the intracellular cAMP concentration. These findings suggest that Gpb1p and Gpb2p mediate their
effects on the cAMP/PKA signaling pathway either by inhibiting the activity of PKA in a cAMP-independent manner or by activating
phosphatases that act on PKA substrates.

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    • "However, recently the existence of a cAMP-independent PKA pathway has begun to emerge in multiple organisms (Blackstone and Chang, 2011; Brown et al., 2013; Graef and Nunnari, 2011; McInnis et al., 2010). In S. cerevisiae the existence of a glucose-sensing cAMP-independent signaling pathway for activation of PKA has been described previously (Budhwar et al., 2010; Lu and Hirsch, 2005 ). Starvation in mammalian cells triggers autophagy, yet mitochondria enlarge in size, which has been shown to sustain ATP production and prevent mitophagy (Blackstone and Chang, 2011). "
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    • "proteins, Krh1 and Krh2, which were also called Gpb2 and Gpb1, referring to a possible role as Gβ subunit for Gpa2 (Harashima & Heitman, 2002). Later work, however, showed that these proteins function in an adenylate cyclase bypass pathway, allowing direct activation of PKA by activated Gpa2 (Lu & Hirsch, 2005, Peeters, et al., 2006). The kelch repeat proteins directly bind to the catalytic subunits of PKA and thereby stimulate association of the catalytic and regulatory subunits of PKA, lowering PKA activity. "
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    • "However, substantial evidence has accumulated discounting Gpb1/Gpb2 as b subunits (Peeters et al. 2007), including the fact that the site on Gpa2 at which the proteins bind does not correspond to the classic Gb-binding domain (Niranjan et al. 2007 ). Nonetheless , Gpb1 and Gpb2 play redundant roles in negatively regulating the activity of the Ras/PKA pathway, either by interference with the Gpr1/Gpa2 interaction (Harashima and Heitman 2005), or through stabilization of the Ras– GAP proteins, Ira1 and Ira2 (Harashima et al. 2006), or by stabilization of the interaction between the regulatory subunit, Bcy1, and the catalytic subunits, Tpk1–3, of protein kinase A (Lu and Hirsch 2005; Peeters et al. 2006; Budhwar et al. 2010), or by some combination of all three mechanisms. One should note that the studies on Gpr1, Gpa2, and Gpb1/2 have not examined the dynamic nature of these components in the context of signal transduction. "
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