The Regulatory Subunit of Protein Kinase A (Bcy1) in Candida albicans Plays Critical Roles in Filamentation and White-Opaque Switching but Is Not Essential for Cell Growth

Abstract

The conserved cAMP-dependent protein kinase (PKA) is composed of the regulatory and catalytic subunits and acts as the central component of the cAMP signaling pathway. In the human fungal pathogen Candida albicans, the PKA regulatory subunit Bcy1 plays a critical role in the regulation of cell differentiation and death. It has long been considered that Bcy1 is essential for cell viability in C. albicans. In the current study, surprisingly, we found that Bcy1 is not required for cell growth, and we successfully generated a bcy1/bcy1 null mutant in C. albicans. Deletion of BCY1 leads to multiple cellular morphologies and promotes the development of filaments. Filamentous and smooth colonies are two typical morphological types of the bcy1/bcy1 mutant, which can undergo spontaneous switching between the two types. Cells of filamentous colonies grow better on a number of different culture media and have a higher survival rate than cells of smooth colonies. In addition, deletion of BCY1 significantly increased the frequency of white-to-opaque switching on N-acetylglucosamine (GlcNAc)-containing medium. The bcy1/bcy1 null mutant generated herein provides the field a new resource to study the biological functions of the cAMP signaling pathway in C. albicans.

Multiple roles and diverse regulation of the Ras/cAMP/protein kinase A pathway in Candida albicans

“…Consistently, the overexpression of RAS1V13 (encoding the active form of Ras1), the overexpression of TPK2 or the deletion of PDE2 causes a mass conversion to the opaque phenotype under the same culture conditions (Huang et al ., ). The deletion of the PKA regulatory subunit‐encoding gene BCY1 also promotes the opaque phenotype and can even induce white‐to‐opaque switching in an MTL a /α background strain, which is normally switching‐incompetent (Ding et al ., ). However, in the presence of Tpk2, the Tpk1 isoform of the PKA catalytic subunit seems to function as a negative regulator of white‐to‐opaque switching in C. albicans (Cao et al ., ).…”

“…Activation of the Ras/cAMP/PKA pathway by overexpressing a constitutively active form of Ras (Ras1V13), the deletion of PDE2 or the deletion of BCY1 promotes filamentation in C. albicans (Feng et al ., ; Jung et al ., ; Ding et al ., ). Consistently, inactivation of this pathway by the deletion of CYR1 or both TPK1 and TPK2 completely blocks the filamentous growth in response to many strong inducers including serum, N‐acetylglucosamine (GlcNAc) and high levels of CO 2 (Klengel et al ., ; Cao et al ., ).…”

“…The Ras/cAMP/PKA pathway has attracted intensive attention in the Candida field in the past two decades. In C. albicans, none of the components of this pathway, including Ras1, Ras2, Cyr1, Pde1, Pde2, Tpk1, Tpk2 and Bcy1, is essential for cell viability, although inactivation of some components results in slow cell growth (Rocha et al, 2001;Jung et al, 2005;Zhu et al, 2009;Wilson et al, 2010;Ding et al, 2016;Cao et al, 2017). The deletion of RAS1 in C. albicans leads to a ~20-fold decrease of intracellular cAMP level, while further deletion of RAS2 restores it to ~30% of the wild-type strain, suggesting that the two Ras proteins are not required for cell viability and play antagonistic roles in the regulation of the cAMP level (Zhu et al, 2009).…”

“…PKA consists of two catalytic and two regulatory subunits and is activated when cAMP binds to the regulatory subunits to release the catalytic subunits. In C. albicans , the PKA catalytic subunits have two isoforms encoded by TPK1 and TPK2 (Bockmuhl et al ., ), whereas the regulatory subunit is encoded by a single gene BCY1 (Giacometti et al ., ; Ding et al ., ). The downstream targets of PKA include transcriptional regulators and other effectors to control cellular responses.…”

“…The major components of the Ras/cAMP/PKA pathway of C. albicans include two Ras GTPases (Ras1 and Ras2) (Feng et al, 1999;Zhu et al, 2009), an adenylyl cyclase (Cyr1, also named as Cdc35) (Rocha et al, 2001), a low (Pde1) and a high (Pde2) affinity cyclic nucleotide phosphodiesterase (Hoyer et al, 1994;Jung and Stateva, 2003) and the PKA comprised of regulatory and catalytic subunits Giacometti et al, 2006;Ding et al, 2016;Cao et al, 2017). C. albicans Ras1 shares high similarity to Ras proteins of other fungi in both the amino acid sequence and domain organization (Feng et al, 1999;Zhu et al, 2009).…”