Pin1 Knockdown Research Articles

Pin1 promotes cell death in NGF‐dependent neurons through a mechanism requiring c‐Jun activity

Developing neurons deprived of trophic support undergo apoptosis mediated by activation of c-Jun N-terminal kinases (JNK) and c-Jun, induction of the Bcl-2 homology 3-only protein Bim(EL), Bax-dependent loss of mitochondrial cytochrome c, and caspase activation. However, the mechanisms that regulate each of these events are only partially understood. Here we show that the prolyl isomerase Pin1 functions as a positive regulator of neuronal death through a c-Jun-dependent mechanism. Ectopic Pin1 promoted caspase-dependent death of NGF-maintained neurons that was associated with an accumulation of Ser(63)-phosphorylated c-Jun in neuronal nuclei and was partially dependent on Bax. Downregulating Pin1 prior to NGF withdrawal suppressed the accumulation of phosphorylated c-Jun, inhibited the release of cytochrome c, and significantly delayed cell death. Pin1 knockdown inhibited NGF deprivation-induced death to a similar extent in Bim (+/+) and Bim (-/-) neurons. The protective effect of Pin1 knockdown was significantly greater than that caused by loss of Bim and nearly identical to that caused by a dominant negative form of c-Jun. Finally, cell death induced by ectopic Pin1 was largely blocked by expression of dominant negative c-Jun. These results suggest a novel mechanism by which Pin1 promotes cell death involving activation of c-Jun.

Pin1 has opposite effects on wild-type and P301L tau stability and tauopathy

Tau pathology is a hallmark of many neurodegenerative diseases including Alzheimer disease (AD) and frontotemporal dementia with Parkinsonism linked to chromosome 17 (FTDP-17). Genetic tau mutations can cause FTDP-17, and mice overexpressing tau mutants such as P301L tau are used as AD models. However, since no tau mutations are found in AD, it remains unclear how appropriate tau mutant mice are as an AD model. The prolyl isomerase Pin1 binds and isomerizes tau and has been implicated in protecting against neurodegeneration, but whether such Pin1 regulation is affected by tau mutations is unknown. Consistent with earlier findings that Pin1 KO induces tauopathy, here we demonstrate that Pin1 knockdown or KO increased WT tau protein stability in vitro and in mice and that Pin1 overexpression suppressed the tauopathy phenotype in WT tau transgenic mice. Unexpectedly, Pin1 knockdown or KO decreased P301L tau protein stability and abolished its robust tauopathy phenotype in mice. In contrast, Pin1 overexpression exacerbated the tauopathy phenotype in P301L tau mice. Thus, Pin1 has opposite effects on the tauopathy phenotype depending on whether the tau is WT or a P301L mutant, indicating the need for disease-specific therapies for tauopathies.

Post‐transcriptional regulation of inducible nitric oxide synthase (iNOS) by Protein Interacting with Never in Mitosis Gene A‐1 (PIN1) in murine aortic endothelial cells

PIN1, a peptidyl‐proline isomerase, increases the level or activity of several transcription factors that can induce iNOS. Thus it was hypothesized that depletion of PIN1 would reduce induction of iNOS by lipopolysaccharide (LPS) and interferon‐gamma (IFN). Murine aortic endothelial cells (EC) were stably transfected with a vector producing a PIN1 short hairpin RNA (shRNA) or an inactive control sequence. PIN1 shRNA reduced PIN1 levels >85% compared to control. Unexpectedly, suppression of PIN 1 enhanced NO production in response to LPS/IFN, and iNOS protein was induced 4.7‐fold more. The increased iNOS was cytotoxic to EC lacking PIN1, but not to control EC, in a crystal violet staining assay. Furthermore, cytotoxicity was prevented by a nitric oxide synthase inhibitor, L‐NAME. LPS/IFN increased iNOS/β‐actin mRNA ratio as expected, but PIN1 knockdown did not affect iNOS mRNA levels measured by RT‐PCR. Instead, PIN1 depletion stabilized iNOS protein in cycloheximide‐treated EC such that the time to a 50% decrease was extended from 8 h to more than 24 h. The loss of iNOS was antagonized by the calpain inhibitor, MDL 28170, but not by the selective proteasome inhibitor, epoxomicin. These results suggest that PIN1 normally restrains the induction of iNOS by facilitating its calpain‐mediated degradation in EC. Modulation of PIN1 may significantly affect iNOS‐dependent vascular activity and inflammatory diseases.