The Not4 E3 Ligase and CCR4 Deadenylase Play Distinct Roles in Protein Quality Control

Abstract

Eukaryotic cells control their proteome by regulating protein production and protein clearance. Protein production is determined to a large extent by mRNA levels, whereas protein degradation depends mostly upon the proteasome. Dysfunction of the proteasome leads to the accumulation of non-functional proteins that can aggregate, be toxic for the cell, and, in extreme cases, lead to cell death. mRNA levels are controlled by their rates of synthesis and degradation. Recent evidence indicates that these rates have oppositely co-evolved to ensure appropriate mRNA levels. This opposite co-evolution has been correlated with the mutations in the Ccr4-Not complex. Consistently, the deadenylation enzymes responsible for the rate-limiting step in eukaryotic mRNA degradation, Caf1 and Ccr4, are subunits of the Ccr4-Not complex. Another subunit of this complex is a RING E3 ligase, Not4. It is essential for cellular protein solubility and has been proposed to be involved in co-translational quality control. An open question has been whether this role of Not4 resides strictly in the regulation of the deadenylation module of the Ccr4-Not complex. However, Not4 is important for proper assembly of the proteasome, and the Ccr4-Not complex may have multiple functional modules that participate in protein quality control in different ways. In this work we studied how the functions of the Caf1/Ccr4 and Not4 modules are connected. We concluded that Not4 plays a role in protein quality control independently of the Ccr4 deadenylase, and that it is involved in clearance of aberrant proteins at least in part via the proteasome.

Translational Capacity of a Cell Is Determined during Transcription Elongation via the Ccr4-Not Complex

“…Recently, we have shown that Not5 plays a role in translatability and assembly of the RNAPII complex (Villanyi et al, 2014). In addition, components of the Ccr4-Not complex, particularly Not4 and Not5, are important for transcription elongation (Kruk et al, 2011) and protein quality control (Dimitrova et al, 2009; Halter et al, 2014; Preissler et al, 2015). Hence these components of the Ccr4-Not complex are prime candidates to coordinate transcription with translation.…”

“…First, analysis of deletion mutants of the subunits of the Ccr4-Not complex revealed that the complex controls expression of most of the genome (Azzouz et al, 2009b; Cui et al, 2008), with particular impacts on snoRNAs (Azzouz et al, 2009a; Halter et al, 2014) and SAGA regulated genes (Cui et al, 2008). Second, genome-wide ChIP experiments revealed the presence of the Ccr4-Not complex on SAGA regulated genes (Venters et al, 2011).…”

Not4‐dependent translational repression is important for cellular protein homeostasis in yeast

“…Not4 contains an N-terminal RING domain required for its ubiquitination activity (Mulder et al, 2007) and has been suggested to regulate the levels of the histone demethylase Jhd2, the catalytic subunit of the DNA polymerase-a Cdc17, the transcription factor Yap1, the nascent polypeptide-associated complex NAC and the ribosomal protein Rps7A (Panasenko et al, 2006;Mersman et al, 2009;Haworth et al, 2010;Gulshan et al, 2012;Panasenko & Collart, 2012). In addition, the deletion of NOT4 affects cellular protein homeostasis (Halter et al, 2014), and based on the observation that the levels of cotranslationally arrested polypeptides were increased in cells lacking Not4, it was proposed that Not4 ubiquitinates arrested nascent polypeptides to target them for proteasomal degradation (Dimitrova et al, 2009). In contrast, other studies suggest that deletion of NOT4 enhances ubiquitination of nascent chains and does not affect the degradation of arrested translation products (Bengtson & Joazeiro, 2010;Duttler et al, 2013).…”

Switch from translation initiation to elongation needs Not4 and Not5 collaboration

“…S6A), released from ribosomes ( Fig. S6B) and new proteins aggregate (21). 15 Based on our observations, we propose that Not5 and Not4 likely regulate the ribosome traffic for Not1-associated mRNAs to avoid downstream ribosomal collisions ( Fig.…”