Protein kinase A regulatory subunit isoforms regulate growth and differentiation in Mucor circinelloides: essential role of PKAR4.
The protein kinase A (PKA) signaling pathway plays a role in regulating growth and differentiation in the dimorphic fungus Mucor circinelloides. PKA holoenzyme is comprised of two catalytic (C) and two regulatory (R) subunits. In M. circinelloides, four genes encode the PKAR1, PKAR2, PKAR3, and PKAR4 isoforms of R subunits. We have constructed null mutants and demonstrate that each isoform has a different role in growth and differentiation. The most striking finding is that pkaR4 is an essential gene, because only heterokaryons were obtained in knockout experiments. Heterokaryons with low levels of wild-type nuclei showed an impediment in the emission of the germ tube, suggesting a pivotal role of this gene in germ tube emergence. The remaining null strains showed different alterations in germ tube emergence, sporulation, and volume of the mother cell. The pkaR2 null mutant showed an accelerated germ tube emission and was the only mutant that germinated under anaerobic conditions when glycine was used as a nitrogen source, suggesting that pkaR2 participates in germ tube emergence by repressing it. From the measurement of the mRNA and protein levels of each isoform in the wild-type and knockout strains, it can be concluded that the expression of each subunit has its own mechanism of differential regulation. The PKAR1 and PKAR2 isoforms are posttranslationally modified by ubiquitylation, suggesting another regulation point in the specificity of the signal transduction. The results indicate that each R isoform has a different role in M. circinelloides physiology, controlling the dimorphism and contributing to the specificity of cyclic AMP (cAMP)-PKA pathway.
Departamento de Química Biológica, Universidad de Buenos Aires, Ciudad Universitaria, Buenos Aires, Argentina.
J
J
Ocampo
B
B
McCormack
E
E
Navarro
S
S
Moreno
V
V
Garre
S
S
Rossi
D017868:Cyclic AMP-Dependent Protein Kinases / Q000235:genetics / Q000378:metabolism*; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D017353:Gene Deletion; D020035:Holoenzymes / Q000235:genetics / Q000378:metabolism; D009024:Morphogenesis / Q000235:genetics; D009089:Mucor / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development*; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D013172:Spores, Fungal / Q000254:growth & development; D054875:Ubiquitination
D005656:Fungal Proteins; D020035:Holoenzymes; D020033:Protein Isoforms; D021122:Protein Subunits; D017868:Cyclic AMP-Dependent Protein Kinases