Abstract
In response to proteasome dysfunction, mammalian cells upregulate proteasome gene expression by activating Nrf1. Nrf1 is an endoplasmic reticulum-resident transcription factor that is continually retrotranslocated and degraded by the proteasome. Upon proteasome inhibition, Nrf1 escapes degradation and is cleaved to become active. However, the processing enzyme for Nrf1 remains obscure. Here we show that the aspartyl protease DNA-damage inducible 1 homolog 2 (DDI2) is required to cleave and activate Nrf1. Deletion of DDI2 reduced the cleaved form of Nrf1 and increased the full-length cytosolic form of Nrf1, resulting in poor upregulation of proteasomes in response to proteasome inhibition. These defects were restored by adding back wild-type DDI2 but not protease-defective DDI2. Our results provide a clue for blocking compensatory proteasome synthesis to improve cancer therapies targeting proteasomes.
Highlights
Proteasome inhibition elicits a response to restore proteasome activity, or a ’bounce-back response,’ where Nrf1 is the responsible transcription factor that upregulates expression of all proteasome subunit genes in a concerted manner in human cells (Radhakrishnan et al, 2010; Steffen et al, 2010)
The ratio of the nuclear to cytoplasmic fluorescent intensities was assessed by high-content microscopy and automated image analysis (Figure 1—figure supplement 1B). p97 small interfering RNA (siRNA) treatment served as a positive control, which abolished Nrf1 translocation following bortezomib treatment while increasing cytoplasmic Nrf1 (Figure 1A) (Radhakrishnan et al, 2014)
The subsequent candidates that had more than two hits in either cell line were examined whether the siRNAs mitigated upregulation of PSMA3, a proteasome subunit gene
Summary
Proteasome inhibition elicits a response to restore proteasome activity, or a ’bounce-back response,’ where Nrf is the responsible transcription factor that upregulates expression of all proteasome subunit genes in a concerted manner in human cells (Radhakrishnan et al, 2010; Steffen et al, 2010). Proteasome inhibitors such as bortezomib and carfilzomib have been in clinical use for treatment of cancers, especially multiple myeloma, but this bounce-back response attenuates the ability of proteasome inhibitors to kill cancer cells (Radhakrishnan et al, 2010). On the other hand, when proteasomes are inhibited, Nrf accumulates and is cleaved into an active form, which moves to the cell nucleus to start producing proteasomes.
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