Plasma membrane calcium ATPase 4b inhibits nitric oxide generation through calcium-induced dynamic interaction with neuronal nitric oxide synthase

The activation and deactivation of Ca^(2+)- and calmodulindependent neuronal nitric oxide synthase (nNOS) in the central nervous system must be tightly controlled to prevent excessive nitric oxide (NO) generation. Considering plasma membrane calcium ATPase (PMCA) is a key deactivator of nNOS, the present investigation aims to determine the key events involved in nNOS deactivation of by PMCA in living cells to maintain its cellular context. Using time-resolved Förster resonance energy transfer (FRET), we determined the occurrence of Ca^(2+)-induced protein-protein interactions between plasma membrane calcium ATPase 4b (PMCA4b) and nNOS in living cells. PMCA activation significantly decreased the intracellular Ca 2+ concentrations ([Ca^(2+)]_(i)), which deactivates nNOS and slowdowns NO synthesis. Under the basal [Ca^(2+)]_(i) caused by PMCA activation, no protein-protein interactions were observed between PMCA4b and nNOS. Furthermore, both the PDZ domain of nNOS and the PDZ-binding motif of PMCA4b were essential for the protein-protein interaction. The involvement of lipid raft microdomains on the activity of PMCA4b and nNOS was also investigated. Unlike other PMCA isoforms, PMCA4 was relatively more concentrated in the raft fractions. Disruption of lipid rafts altered the intracellular localization of PMCA4b and affected the interaction between PMCA4b and nNOS, which suggest that the unique lipid raft distribution of PMCA4 may be responsible for its regulation of nNOS activity. In summary, lipid rafts may act as platforms for the PMCA4b regulation of nNOS activity and the transient tethering of nNOS to PMCA4b is responsible for rapid nNOS deactivation.

Lysosomal chymotrypsin induces mitochondrial fission in apoptotic cells by proteolytic activation of calcineurin

Dear Editor Apoptosis is a fundamental physiological process in mam- mals in which cells die by activating a suicide mechanism. The mitochondria are one of the major checkpoints in apoptotic regulation because they serve as sensors and amplifiers of cellular damage （Green and Kroemer, 2004）. After mitochondrial outer membrane permeabilization （MOMP）, the mitochondria release a number of factors that are critically involved in cell death signaling （Tait and Green, 2010）. Bcl-2 family members are regarded as the key reg- ulators of mitochondria-dependent apoptosis （Moldoveanu et al., 2014）; however, dynamin-related protein 1 （Drpl）, which orchestrates mitochondrial fission, also participates in apoptotic regulation by stimulating Bax oligomerization and thereby enhances MOMP （Montessuit et al., 2010）; accord- ingly, the inhibition of Drpl blocks mitochondrial fission and inhibits apoptosis （Cassidy-Stone et al., 2008）.

Surface preparation by mechanical polishing of the 1.3-GHzmono-cell copper cavity substrate prior chemical etching for niobium coating

Purpose Surface quality of the substrate is widely acknowledged to be essential for the niobium thin film deposition. Mucheffort has thus been spent to improve the surface roughness by using various chemical etching techniques. However, surfacepreparation before the chemical etching also plays a part in obtaining a satisfactory substrate, but has rarely been studiedbefore. This paper aims to define a specification for the pre-polished copper substrate prior chemical etching and searches forsuitable alternative non-chemical grinding methods for the copper cavity.Methods Copper samples were mechanically pre-polished at first by using flap sanding wheels of different grits and thenchemically etched by using the well-established SUBU solutions. Surface roughness, as a figure of merit, was measuredand compared before and after SUBU. Optimum practice for pre-polishing may therefore be determined. The mechanicalgrinding was subsequently applied on the 1.3-GHz mono-cell copper cavity. Meantime, the previously reported centrifugalbarrel polishing method was also applied with new abrasive materials and modified schemes. A comprehensive study ofetching rate, surface roughness and morphologies was conducted.Results and conclusions The specification for surface roughness prior SUBU was determined. Due to a complex geometryand curved surfaces possessed by the 1.3-GHz copper cavity, the traditional mechanical grinding was proved to be not ideal.Satisfactory surface quality was obtained by using the alternative centrifugal barrel polishing on the cavity. The proposed newscheme and new abrasive materials were demonstrated to be effective, and a mirror-like surface was achieved on the coppercavity. The traditional mechanical grinding can therefore be replaced. This constitutes a dedicated study on pre-polishing ofthe 1.3-GHz copper cavity substrate prior chemical etching for niobium sputtering.

OFHC copper substrates for niobium sputtering:comparison of chemical etching recipes

Purpose Niobium sputtered on copper has been a popular alternative approach for superconducting radio frequency(SRF)community in the last few decades.Comparing to bulk materials of a few millimeters,high-purity niobium of merely a few microns is sufficient to realize superconductivity on the coated surface.Being niobium thin film,it has been widely acknowledged that surface quality of the substrate plays a vital role in obtaining a superior niobium coating with excellent SRF performance.Therefore,proper chemical treatment of the substrate before coating is crucial and the ultimate goal is to create a smooth and defect-free surface.Prior to the design of a cavity etching system,the mechanism of SUBU as well as two industry-used solutions is studied in detail on samples.Methods Copper samples were first pre-treated by mechanical grinding to remove fabrication damages,obvious defects and visible impurities.Two chemical solutions widely used in industries were subsequently chosen to etch the samples.Finally,the established SUBU solution was used independently on these pre-treated samples for comparison.Surface morphology and etching rate were measured accordingly.Results and conclusions Mirror-like copper surface was created by using the SUBU solution thus qualified for subsequent niobium sputtering,while the other two solutions used in industries were less effective with nonideal surface morphology.The chemical reactions,the experimental requisites and the involved processes are extensively elucidated for all three solutions.Limitations for SUBU were examined,and the optimum ratio of the chemical bath volume to sample surface area was also determined.These investigations will serve as an important guidance for the development of a chemical etching system for elliptical copper cavities.