A yeast culture grown in a nutrient-rich medium initially containing 2% glucose is not limited in calorie supply. When yeast cells cultured in this medium consume glucose, they undergo cell cycle arrest at a checkpoint in late G1 and differentiate into quiescent and non-quiescent cell populations. Studies of such differentiation have provided insights into mechanisms of yeast chronological aging under conditions of excessive calorie intake. Caloric restriction is an aging-delaying dietary intervention. Here, we assessed how caloric restriction influences the differentiation of chronologically aging yeast cultures into quiescent and non-quiescent cells, and how it affects their properties. We found that caloric restriction extends yeast chronological lifespan via a mechanism linking cellular aging to cell cycle regulation, maintenance of quiescence, entry into a non-quiescent state and survival in this state. Our findings suggest that caloric restriction delays yeast chronological aging by causing specific changes in the following: 1) a checkpoint in G1 for cell cycle arrest and entry into a quiescent state; 2) a growth phase in which high-density quiescent cells are committed to become low-density quiescent cells; 3) the differentiation of low-density quiescent cells into low-density non-quiescent cells; and 4) the conversion of high-density quiescent cells into high-density non-quiescent cells.

Eukaryotic mRNA degradation often initiates with the recruitment of the CCR4-NOT deadenylase complex and decay factors to the mRNA 3' terminus. How the 3'-proximal decay machinery interacts with the 5'-terminal cap structure in order to engender mRNA decapping and 5'-3' degradation is unclear. Human 4E-T is an eIF4E-binding protein that has been reported to promote mRNA decay, albeit via an unknown mechanism. Here, we show that 4E-T is a component of the mRNA decay machinery and interacts with factors including DDX6, LSM14, and the LSM1-7-PAT1 complex. We also provide evidence that 4E-T associates with, and enhances the decay of, mRNAs targeted by the CCR4-NOT deadenylase complex, including microRNA targets. Importantly, we demonstrate that 4E-T must interact with eIF4E to engender mRNA decay. Taken together, our data support a model where 4E-T promotes mRNA turnover by physically linking the 3'-terminal mRNA decay machinery to the 5' cap via its interaction with eIF4E.

The LSM domain-containing protein LSM14/Rap55 plays a role in mRNA decapping, translational repression, and RNA granule (P-body) assembly. How LSM14 interacts with the mRNA silencing machinery, including the eIF4E-binding protein 4E-T and the DEAD-box helicase DDX6, is poorly understood. Here we report the crystal structure of the LSM domain of LSM14 bound to a highly conserved C-terminal fragment of 4E-T. The 4E-T C-terminus forms a bi-partite motif that wraps around the N-terminal LSM domain of LSM14. We also determined the crystal structure of LSM14 bound to the C-terminal RecA-like domain of DDX6. LSM14 binds DDX6 via a unique non-contiguous motif with distinct directionality as compared to other DDX6-interacting proteins. Together with mutational and proteomic studies, the LSM14-DDX6 structure reveals that LSM14 has adopted a divergent mode of binding DDX6 in order to support the formation of mRNA silencing complexes and P-body assembly.