Analysis of All Protein Phosphatase Genes in Aspergillus nidulans Identifies a New Mitotic Regulator, Fcp1

Abstract

Reversible protein phosphorylation is an important regulatory mechanism of cell cycle control in which protein phosphatases counteract the activities of protein kinases. In Aspergillus nidulans, 28 protein phosphatase catalytic subunit genes were identified. Systematic deletion analysis identified four essential phosphatases and four required for normal growth. Conditional alleles of these were generated using the alcA promoter. The deleted phosphatase strain collection and regulatable versions of the essential and nearessential phosphatases provide an important resource for further analysis of the role of reversible protein phosphorylation to the biology of A. nidulans. We further demonstrate that nimT and bimG have essential functions required for mitotic progression since their deletions led to classical G 2 -and M-phase arrest. Although not as obvious, cells with AnpphA and Annem1 deleted also have mitotic abnormalities. One of the essential phosphatases, the RNA polymerase II C-terminal domain phosphatase Anfcp1, was further examined for potential functions in mitosis because a temperature-sensitive Anfcp1 allele was isolated in a genetic screen showing synthetic interaction with the cdk1F mutation, a hyperactive mitotic kinase. The Anfcp1 ts cdk1F double mutant had severe mitotic defects, including inability of nuclei to complete mitosis in a normal fashion. The severity of the Anfcp1 ts cdk1F mitotic phenotypes were far greater than either single mutant, confirming the synthetic nature of their genetic interaction. The mitotic defects of the Anfcp1 ts cdk1F double mutant suggests a previously unrealized function for AnFCP1 in regulating mitotic progression, perhaps counteracting Cdk1-mediated phosphorylation.In many cellular processes reversible phosphorylation/dephosphorylation reactions play important roles by regulating protein activity and subcellular localization. As enzymes counteracting the function of protein kinases, protein phosphatases have been firmly established as key coordinators of diverse biological events. Three main criteria are used to classify protein phosphatases: sequence homology, structure, and catalytic mechanism concerning substrate specificity. According to these features, protein phosphatases are divided into the three main groups: classical serine/threonine phosphatases, protein tyrosine phosphatases, and the aspartate based catalysis protein phosphatase (reviewed in reference 33).The classical Ser/Thr phosphatases can be further subdivided into the phosphoprotein phosphatase (PPP) family and the protein phosphatase Mg 2ϩ /Mn 2ϩ -dependent (PPM) family. All members of the PPP family share high sequence similarities and have in common a conserved phosphoesterase catalytic motif (PP2Ac), although their biological functions range widely from mitotic regulation (PP1) (8), immune response (PP2B) (4), DNA damage response (PP4) (15), and blue-light signaling in plants (PP7) (32). PP2C type protein phosphatases make up the PPM family and are characterized by structural similarity of ...

Functional Characterization of a New Member of the Cdk9 Family in Aspergillus nidulans

“…Since A. nidulans undergoes a partially open mitosis (6, 7), this indicates that PtkA does not remain associated with nuclear structures during nuclear division. A similar localization pattern was also observed for RNAPII (47), possibly reflecting the general downregulation of transcription during mitosis (57,58).…”

Type 2C Protein Phosphatases in Fungi

“…16 in Fig. S1 in the supplemental (93), although no further characterization was carried out. It must be noted that according to our analysis (see above), this organism does not contain two Ptc2 isoforms, as proposed by these authors.…”

Protein Phosphatase 2A (PP2A) Regulatory Subunits ParA and PabA Orchestrate Septation and Conidiation and Are Essential for PP2A Activity in Aspergillus nidulans

“…To date, 28 protein phosphatase catalytic subunit genes have been identified in A. nidulans (36). Among them, a gene for the catalytic subunit of PP2A (pphA) is able to regulate hyphal morphogenesis, and a site-mutated mutant form of pphA (R259/Q) leads to slow growth, delayed germ tube emergence, and mitotic defects at low temperatures (37).…”