Tmc1 Is a Dynamically Regulated Effector of the Rpn4 Proteotoxic Stress Response

Abstract

The ubiquitin-proteasome system represents the major pathway of selective intracellular protein degradation in eukaryotes. Misfolded proteins represent an important class of substrates for this pathway, and the failure to destroy misfolded proteins is associated with a number of human diseases. The transcription factor Rpn4 mediates a key proteotoxic stress response whose best known function is to control proteasome abundance by a homeostatic feedback mechanism. Here we identify the uncharacterized zinc finger protein Tmc1 as a dynamically regulated stress-responsive protein. Rpn4 induces TMC1 transcription in response to misfolded proteins. However, this response is counteracted by rapid proteasome-dependent degradation of Tmc1, which serves to normalize Tmc1 protein levels after induction. Precise control of Tmc1 levels is needed in vivo to survive multiple stressors related to proteostasis. Thus, Tmc1 represents a novel effector and substrate of the Rpn4 proteotoxic stress response.

Induction of proteotoxic stress by the mycotoxin patulin

“…Central to the cellular response to arsenic were broad changes in pathways related to protein homeostasis, or proteostasis, including regulation of both protein synthesis and protein degradation. In particular we observed an approximately 20-fold upregulation of Rpn4 at the protein level at one hour after arsenic treatment, and this was accompanied by an increase in the protein levels of most proteasome subunits (Guerra-Moreno et al, 2015; Guerra-Moreno et al, 2016). This upregulation of Rpn4 was consistent with a response to proteotoxic stress, and appeared to be efficacious inasmuch as Rpn4 levels, after initially rising, tended to normalize over time.…”

“…Reverse transcription-PCR was performed as described previously (Guerra-Moreno et al, 2016; Weisshaar et al, 2017). The following oligonucleotides were used to amplify RPN4 : 5-CTAATCCAGGTCACAGTCAG-3 and 5-GGTACATACGGGTCTGATAC-3.…”

Targeted Degradation of Glucose Transporters Protects against Arsenic Toxicity

“…The efficacy of proteasome inhibition was confirmed by the strong accumulation of highmolecular-weight ubiquitin-immunoreactive material as well as a specific proteasome substrate, Tmc1 ( Fig. 2B; compare the first and third lanes) (24). In contrast, Hxt7 downregulation was completely unaffected by proteasome inhibition (Fig.…”

“…We sought to determine the E2 enzyme for this process. In previous work, we had determined the arsenite sensitivities of seven E2 null mutants (mutants lacking rad6 and ubc4, -5, -7, -8, -11, and -13) (24). Only the ubc4Δ mutant showed a strong growth defect upon arsenic treatment, suggesting an important role for Ubc4 in the response to arsenic (24).…”

“…In previous work, we had determined the arsenite sensitivities of seven E2 null mutants (mutants lacking rad6 and ubc4, -5, -7, -8, -11, and -13) (24). Only the ubc4Δ mutant showed a strong growth defect upon arsenic treatment, suggesting an important role for Ubc4 in the response to arsenic (24). Interestingly, Ubc5, which is highly similar to Ubc4 at the amino acid level (29), showed no such phenotype.…”