Background: PTEN is targeted by multiple E3 ligases but that does not clearly decipher the rigid control of its level and activity. Results: CHIP interacts with PTEN and promotes its proteasomal degradation. Conclusion: CHIP acts as a bridge between the chaperone system and the degradation machinery for PTEN. Significance: Stabilization of PTEN by targeting CHIP can be a novel therapeutic approach in cancer regulation.

c-Myc is a proto-oncogenic transcription factor and its rapid turnover mediated by the ubiquitin-proteasome system is critical for maintaining normal cellular homeostasis. Multiple ubiquitin ligases have been assigned for c-Myc regulation till date. However, the available data suggest for the possible existence of additional E3 ligase(s). Here, we report a new E3 ligase for c-Myc, the carboxyl terminus of Hsc70-interacting protein or CHIP, which is a chaperone-associated Ubox-containing E3 ligase. In this report, we show that CHIP interacts and ubiquitinates c-Myc, thus targeting it for proteasome-mediated degradation. Overexpression of CHIP could accelerate the turnover rate of c-Myc protein. Conversely, knockdown of CHIP by RNAi stabilizes endogenous c-Myc. The interaction between CHIP and c-Myc depends on the N-terminally located tetratricopeptide repeats of CHIP, which has been implicated as a chaperone-binding motif. Inhibition of Hsp90 chaperone activity by 17-N-allylamino-17-demethoxygeldanamycin reduces c-Myc protein level. We found that the association between CHIP and c-Myc is dependent on the chaperones; particularly Hsp70. CHIP antagonizes the transcriptional activity of c-Myc and decreases the abundance of the transcripts of its target genes. Overall, CHIP-knockdown increases malignant behavior of C6 glioma cells. To the best of our knowledge, this is the first report of c-Myc being regulated by a bona-fide chaperone-associated E3 ligase in HEK293 as well as glioma cells. Because CHIP has been reported earlier to be negatively regulating Akt1, BCR-ABL and hTERT, and now c-Myc, the present study may strengthen the view that CHIP acts as a tumor suppressor.

Strain ST-14, characterized as a member of the genus Cupriavidus, was capable of utilizing 2-and 4-nitrobenzoates individually as sole sources of carbon and energy. Biochemical studies revealed the assimilation of 2-and 4-nitrobenzoates via 3-hydroxyanthranilate and protocatechuate, respectively. Screening of a genomic fosmid library of strain ST-14 constructed in Escherichia coli identified two gene clusters, onb and pob-pca, to be responsible for the complete degradation of 2-nitrobenzoate and protocatechuate, respectively. Additionally, a gene segment (pnb) harboring the genes for the conversion of 4-nitrobenzoate to protocatechuate was unveiled by transposome mutagenesis. Reverse transcription-PCR analysis showed the polycistronic nature of the gene clusters, and their importance in the degradation of 2-and 4-nitrobenzoates was ascertained by gene knockout analysis. Cloning and expression of the relevant pathway genes revealed the transformation of 2-nitrobenzoate to 3-hydroxyanthranilate and of 4-nitrobenzoate to protocatechuate. Finally, incorporation of functional 3-nitrobenzoate dioxygenase into strain ST-14 allowed the recombinant strain to utilize 3-nitrobenzoate via the existing protocatechuate metabolic pathway, thereby allowing the degradation of all three isomers of mononitrobenzoate by a single bacterial strain. IMPORTANCEMononitrobenzoates are toxic chemicals largely used for the production of various value-added products and enter the ecosystem through industrial wastes. Bacteria capable of degrading mononitrobenzoates are relatively limited. Unlike other contaminants, these man-made chemicals have entered the environment since the last century, and it is believed that bacteria in nature evolved not quite efficiently to assimilate these compounds; as a consequence, to date, there are only a few reports on the bacterial degradation of one or more isomers of mononitrobenzoate. In the present study, fortunately, we have been able to isolate a Cupriavidus sp. strain capable of assimilating both 2-and 4-nitrobenzoates as the sole carbon source. Results of the biochemical and molecular characterization of catabolic genes responsible for the degradation of mononitrobenzoates led us to manipulate a single enzymatic step, allowing the recombinant host organism to expand its catabolic potential to assimilate 3-nitrobenzoate.