capturing-interaction-kinetics-an-ion-channel-protein-with-small

Capturing the Interaction Kinetics of an Ion Channel Protein with Small Molecules by the Bio-layer Interferometry Assay

The bio-layer interferometry (BLI) assay is a valuable tool for measuring protein-protein and protein-small molecule interactions. Here, we first describe ...

Shanghai Jiao Tong University

c-Abl is a proto-oncogene that is essential for mouse development and tissue homeostasis. Misregulation of c-Abl, as seen in the constitutively active BCR-ABL, is the leading cause of human chronic myeloid leukemia. However, how the Abl proteins execute their functions still remains largely unknown. Here, we report an important role for c-Abl in replicative senescence and immortalization by regulating the expression of two tumor suppressors that induce cellular senescence, p53 and p16(INK4a). Using primary mouse embryonic fibroblasts (MEFs), we show that c-Abl (-/-) cells were more resistant to immortalization than wildtype cells using a standard 3T3 or 3T9 protocol. We could only immortalize three out of nine c-Abl (-/-) MEF cultures even when we increased the number of starting cells. This resistance was attributed to premature senescence and reduced survival in senescent c-Abl (-/-) cells due to an increase in p16(INK4a) and p53 expression. Deleting p53 allows c-Abl (-/-) p53 (-/-) MEFs to bypass senescence to be spontaneously immortalized. Cell immortalization, but not senescence, was generally accompanied by mutations in p53 in both wildtype and c-Abl (-/-) MEFs, although the spectrum is different from that of human tumors. The role for c-Abl in regulating cell senescence and immortalization might explain some of the developmental defects in c-Abl (-/-) mice and how BCR-ABL transforms cells.

A role for c-Abl in cell senescence and spontaneous immortalization.

C-kit positive (c-kit(+)) cells are usual tissue-specific stem cells. However, in postnatal testis, undifferentiated spermatogonial stem cells (SSCs) are c-kit negative (c-kit(-)) and activation of c-kit represents the start of SSC differentiation, leaving an intriguing question whether other c-kit(+) cells exist and participate in the postnatal development of testis. To this end, a feasible system for testicular reconstitution, in which a specific type of cells can be manipulated, is needed. Here, we first establish de novo morphogenesis of testis by subcutaneous injection of testicular cells from neonatal testes into the backs of nude mice. We observe testicular tissue formation and spermatogenesis from all injected sites. Importantly, functional spermatids can be isolated from these testicular tissues. Using this system, we systemically analyze the roles of c-kit(+) cells in testicular reconstitution and identify a small population of cells (c-kit(+):CD140a(+):F4/80(+)), which express typical markers of macrophages, are critical for de novo morphogenesis of testis. Interestingly, we demonstrate that these cells are gradually replaced by peripheral blood cells of recipient mice during the morphogenesis of testis. Thus, we develop a system, which may mimic the complete developmental process of postnatal testis, for investigating the testicular development and spermatogenesis.

The roles of testicular c-kit positive cells in de novo morphogenesis of testis.

N-nitrosodimethylamine (NDMA) photocatalytic degradation performance and mechanism were investigated on the TiO2 nanotube prepared from anatase TiO2 nanopowder in terms of the production of reactive oxygen species including hydroxyl radical, singlet oxygen and superoxide radical. Significantly higher NDMA degradation efficiency was obtained on anatase TiO2 nanotube rather than anatase TiO2 nanopowder. The tubular morphology may be responsible for almost 100% NDMA removal on TiO2 nanotube, presumably due to its confinement effect leading to NDMA molecules within the nanotube being attacked by reactive oxygen species such as hydroxyl radical and singlet oxygen, and initiating reaction inside the nanotube. In particular, the ability of the nanotubular structure of TiO2 nanotube to promote a singlet oxygen oxidation pathway contributes much to the enhanced NDMA degradation efficiency and favors the formation of dimethylamine and NO3(-). Such function originating from nanotube morphology could bring new insights for the photocatalytic degradation of organic pollutants.

Enhanced photocatalytic performance of N-nitrosodimethylamine on TiO2 nanotube based on the role of singlet oxygen.

A series of CAPE derivatives with mono-substituted phenylethanols moiety were synthesized and evaluated by MTT assay on growth of 4 human cancer cell lines (Hela, DU-145, MCF-7 and ECA-109). The substituent effects on the antiproliferative activity were systematically investigated for the first time. It was found that electron-donating and hydrophobic substituents at 2'-position of phenylethanol moiety could significantly enhance CAPE's antiproliferative activity. 2'-Propoxyl derivative, as a novel caffeic acid ester, exhibited exquisite potency (IC=0.4±0.02 & 0.6±0.03μM against Hela and DU-145 respectively).

Antiproliferative activity and SARs of caffeic acid esters with mono-substituted phenylethanols moiety.

Genetic studies have elucidated critical roles of Piwi proteins in germline development in animals, but whether Piwi is an actual disease gene in human infertility remains unknown. We report germline mutations in human Piwi (Hiwi) in patients with azoospermia that prevent its ubiquitination and degradation. By modeling such mutations in Piwi (Miwi) knockin mice, we demonstrate that the genetic defects are directly responsible for male infertility. Mechanistically, we show that MIWI binds the histone ubiquitin ligase RNF8 in a Piwi-interacting RNA (piRNA)-independent manner, and MIWI stabilization sequesters RNF8 in the cytoplasm of late spermatids. The resulting aberrant sperm show histone retention, abnormal morphology, and severely compromised activity, which can be functionally rescued via blocking RNF8-MIWI interaction in spermatids with an RNF8-N peptide. Collectively, our findings identify Piwi as a factor in human infertility and reveal its role in regulating the histone-to-protamine exchange during spermiogenesis.

Article	Year
A role for c-Abl in cell senescence and spontaneous immortalization. Age (Dordrecht, Netherlands)\| PubMed ID: 22791394	2013
The roles of testicular c-kit positive cells in de novo morphogenesis of testis. Scientific reports\| PubMed ID: 25088917	2014
Enhanced photocatalytic performance of N-nitrosodimethylamine on TiO2 nanotube based on the role of singlet oxygen. Chemosphere\| PubMed ID: 25290358	2015
Antiproliferative activity and SARs of caffeic acid esters with mono-substituted phenylethanols moiety. Bioorganic & medicinal chemistry letters\| PubMed ID: 27979593	2017
Ubiquitination-Deficient Mutations in Human Piwi Cause Male Infertility by Impairing Histone-to-Protamine Exchange during Spermiogenesis. Cell\| PubMed ID: 28552346	2017

Man Zhang

Biography

JoVE Journal Publications

Other Publications