High impact information on NET1

• During anaphase, Cdk- and Polo-dependent phosphorylation of Net1 is thought to release active Cdc14 [1].
• The sister chromatid-separating protease separase, activated at anaphase onset, interacts with and downregulates PP2A(Cdc55), thereby facilitating Cdk-dependent Net1 phosphorylation [1].
• How Net1 is phosphorylated specifically in anaphase, when mitotic kinase activity starts to decline, has remained unexplained [1].
• Here, we show that PP2A(Cdc55) phosphatase keeps Net1 underphosphorylated in metaphase [1].
• Net1 is furthermore required for rDNA silencing and nucleolar integrity [2].

Biological context of NET1

• NET1 and HFI1 genes of yeast mediate both chromosome maintenance and mitochondrial rho(-) mutagenesis [3].
• In budding yeast, the phosphatase Cdc14, a key regulator of exit from mitosis, is released from its inhibitor Cfi1/Net1 in the nucleolus during anaphase [4].
• Cdc14 is sequestered in the nucleolus by its inhibitor Net1/Cfi1 and is only released from the nucleolus during anaphase to inactivate mitotic CDK [5].
• Here we show that Cdc5 affects the phosphorylation state of Net1 [6].
• During interphase, Net1 and Sir2 colocalize to a subdomain within the nucleous, but at the end of mitosis a fraction of Sir2 leaves the nucleolus and disperses as foci throughout the nucleus, suggesting that the structure of rDNA silent chromatin changes during the cell cycle [2].

Associations of NET1 with chemical compounds

• We report here the phosphorylation site mapping of recombinant Net1 (Net1N) and a mutant Net1N allele (Net1N-19m) with 19 serines or threonines mutated to alanine [7].

Physical interactions of NET1

• An analysis of truncation mutants indicates that the Cdc14 binding site is located within a segment of Net1 containing residues 1-341 [8].

Enzymatic interactions of NET1

• Budding yeast Cdc5 phosphorylates Net1 and assists Cdc14 release from the nucleolus [6].

Regulatory relationships of NET1

• From G1 through anaphase, RENT localizes to the nucleolus, and Cdc14 activity is inhibited by Net1 [9].
• Cdc5 influences phosphorylation of Net1 and disassembly of the RENT complex [10].
• Net1 stimulates RNA polymerase I transcription and regulates nucleolar structure independently of controlling mitotic exit [11].

Other interactions of NET1

• These two genes are NET1 and HFI1 which mediate important regulatory protein-protein interactions in the yeast cell [3].
• Several rDNA silencing mutants display a Sir2p nucleolar localization defect that correlates with reduced Net1p binding [12].
• Furthermore, the Fob1-dependent associations of Net1 and Sir2 with the recombination hotspot region strongly suggest that Sir2 acts directly at this region to carry out its inhibitory effect on rDNA recombination and accelerated aging [13].
• In agreement with this, Net1 provided ORC-dependent, Sir1-independent silencing when artificially tethered to the silencer [14].

References