Selective degradation of PL2L60 by metabolic stresses­induced autophagy suppresses multi­cancer growth.

It has been reported that PL2L60 proteins, a product of PIWIL2 gene which might be activated by an intragenic promoter, could mediate a common pathway specifically for tumorigenesis. In the present study, it was further identified by using western blot assay that the PL2L60 proteins could be degraded in cancer cells through a mechanism of selective autophagy in response to oxidative stress. The PL2L60 was downregulated in various types of cancer cells under the hypoxic condition independently of HIF­1α, resulting in apoptosis of cancer cells. Inhibition of autophagy by small interfering RNA targeting of either Beclin­1 (BECN1) or Atg5 resulted in restoration of PL2L60 expression in hypoxic cancer cell. The hypoxic degradation of PL2L60 was also blocked by the attenuation of the autophagosome membrane protein Atg8/microtubule­associated protein 1 light chain 3 (LC3) or autophagy cargo protein p62 expression. Surprisingly, Immunofluorescence analysis demonstrated that LC3 could be directly bound to PL2L60 and was required for the transport of PL2L60 from the nucleus to the cytoplasm for lysosomal flux under basal or activated autophagy in cancer cells. Moreover, flow cytometric analysis displayed that knocking down of PL2L60 mRNA but not PIWIL2 mRNA effectively inhibited cancer cell proliferation and promoted apoptosis of cancer cells. The similar results were obtained from in vivo tumorigenic experiment, in which PL2L60 downregulation in necroptosis areas was confirmed by immunohistochemistry. These results suggested that various cancer could be suppressed by promoting autophagy. The present study revealed a key role of autophagic degradation of PL2L60 in hypoxia­induced cancer cell death, which could be used as a novel therapeutic target of cancer.

GIT1 enhances neurite outgrowth by stimulating microtubule assembly.

GIT1, a G-protein-coupled receptor kinase interacting protein, has been reported to be involved in neurite outgrowth. However, the neurobiological functions of the protein remain unclear. In this study, we found that GIT1 was highly expressed in the nervous system, and its expression was maintained throughout all stages of neuritogenesis in the brain. In primary cultured mouse hippocampal neurons from GIT1 knockout mice, there was a significant reduction in total neurite length per neuron, as well as in the average length of axon-like structures, which could not be prevented by nerve growth factor treatment. Overexpression of GIT1 significantly promoted axon growth and fully rescued the axon outgrowth defect in the primary hippocampal neuron cultures from GIT1 knockout mice. The GIT1 N terminal region, including the ADP ribosylation factor-GTPase activating protein domain, the ankyrin domains and the Spa2 homology domain, were sufficient to enhance axonal extension. Importantly, GIT1 bound to many tubulin proteins and microtubule-associated proteins, and it accelerated microtubule assembly in vitro. Collectively, our findings suggest that GIT1 promotes neurite outgrowth, at least partially by stimulating microtubule assembly. This study provides new insight into the cellular and molecular pathogenesis of GIT1-associated neurological diseases.

OIT3 deficiency impairs uric acid reabsorption in renal tubule.

The oncoprotein induced transcript 3 (OIT3), also named liver-specific zona pellucida domain-containing protein (LZP), has been shown to be expressed in kidney, and was confirmed to interact with the Tamm-Horsfall glycoprotein (THP). However, the function of OIT3 in kidney remains unclear. In this study we found that serum uric acid level of Oit3 null mice was significantly lower than that in wild type controls, whereas the excretion of uric acid in urine increased in the mutant mouse. Significantly, the excretion of THP in urine also increased while renal THP decreased in Oit3 null mice. Our data suggest that OIT3 could maintain urate homeostasis by regulating the excretion and reabsorption of uric acid in renal tubule via cooperating with THP.

Insulin receptor tyrosine kinase substrate enhances low levels of MDM2-mediated p53 ubiquitination.

The tumor suppressor p53 controls multiple cellular functions including DNA repair, cell cycle arrest and apoptosis. MDM2-mediated p53 ubiquitination affects both degradation and cytoplasmic localization of p53. Several cofactors are known to modulate MDM2-mediated p53 ubiquitination and proteasomal degradation. Here we show that IRTKS, a novel IRSp53-like protein inhibited p53-induced apoptosis and depressed its transcription activity. IRTKS bound directly to p53 and increased p53 ubiquitination and cytoplasmic localization. Further studies revealed that IRTKS interacted with MDM2 and promoted low levels of MDM2-mediated p53 ubiquitination in vitro and in vivo. In unstressed cells with low levels of MDM2, IRTKS was found to stabilize the interaction of p53 and MDM2. In stressed cells, IRTKS dissociated from p53, and high levels of MDM2 induced by p53 activation mediate IRTKS poly-ubiquitination and subsequent proteasomal degradation. These data suggest that IRTKS is a novel regulator of p53, modulating low level of MDM2-mediated p53 ubiquitination in unstressed cells.

Liver-specific ZP domain-containing protein (LZP) as a new partner of Tamm-Horsfall protein harbors on renal tubules.

Liver-specific ZP domain-containing protein (LZP) was recently identified as a secreted protein that is specifically expressed in liver. However, the physiological effects of LZP are largely unknown. In this study, we found that LZP was detectable in mouse kidneys, testes, ovaries and heart, in addition to liver. LZP was localized in the spermatid cells of testes, corpus luteum cells of ovaries, and cardiac muscle cells of heart. But the protein mainly anchored on the apical membrane of the thick ascending limb of the loop of Henle (TAL) cell in mouse kidney. In rat kidney LZP and Tamm-Horsfall protein (THP) were co-localized in TAL. The in vivo interaction between LZP and THP was confirmed in kidney and urine by co-immunoprecipitation assay, and the in vitro interaction was detected by GST pull-down assay, implying that the interaction could be independent on N-linked glycosylated modification of LZP. Surprisingly, LZPs with intramolecular disulfide bridges could self-interact, and then self-aggregate into spheres of varying sizes, but not polymerize into filaments. The finding that LZP might act as a new partner of THP would provide novel insights into renal functions related to THP and LZP, such as the urothelial permeability barrier and the host defense against the adhesion of pathogens.