In the absence of external signals, yeast enter the cell cycle once they have achieved a critical size, and this is controlled by the G1 cyclin Cln3 and its partner, the cyclin-dependent kinase Cdc28. Vergés et al. found that a population of Cln3 and Cdc28 was associated with the cytosolic side of the endoplasmic reticulum (ER) and that the abundance of these proteins in this fraction was highest in "newly born" cells and less in cells in late G1. Cln3 contains a domain, the J domain, like that found in regulators of heat shock protein 70 (HSP70) chaperones such as Ydj1; however, the Cln3 J domain lacked key residues required for adenosine triphosphatase (ATPase) activation of HSP70 chaperones. Cells in which the Cln3 J domain was deleted or mutated were larger, and these mutant versions of Cln3 did not associate with the ER and accumulated in the nucleus. Although Cdc28 was associated with the ER in early G1 cells, a Cln3 mutant that could not interact with Cdc28 accumulated in the nucleus, suggesting that the interaction with Cdc28 was necessary for ER retention. The J chaperone Ydj1, which activates the ATPase activity of the HSP70 chaperones Ssa1 and Ssa2, interacts with Cln3 and promotes Cln3 phosphorylation and degradation. However, deficiencies in Ydj1 or its HSP70 partners Ssa1 and Ssa2 lead to cells with a larger than normal size, and Cln3 does not accumulate in the nucleus of these cells. Chimeric proteins in which the J domains had been swapped between Cln3 and Ydj1 showed that each of these J domains interacted with Ssa1, but only the Ydj1 J domain stimulated ATPase activity. Overexpression of Ydj1 and Ssa1 caused Cln3 to accumulate in the nucleus, and the cells budded at smaller cell sizes than did wild-type cells; premature cell division did not occur in cells deficient for Cln3. In a cell-free assay, excess Ydj1 and Ssa1 released Cln3. The authors suggest that as cells grow and Ydj1 becomes more abundant, it competes with Cln3 for interaction with Ssa1 at the ER, thereby promoting release of Cln3 and Cdc28 to translocate to the nucleus and stimulate entry into the cell cycle.
E. Vergés, N. Colomina, E. Garí, C. Gallego, M. Aldea, Cycline Cln3 is retained at the ER and released by the J chaperone Ydj1 in late G1 to trigger cell cycle entry. Mol. Cell 26, 649-662 (2007). [Online Journal]