Kdm3a lysine demethylase is an Hsp90 client required for cytoskeletal rearrangements during spermatogenesis

The lysine demethylase Kdm3a (Jhdm2a, Jmjd1a) is required for male fertility, sex determination, and metabolic homeostasis through its nuclear role in chromatin remodeling. Many histone-modifying enzymes have additional nonhistone substrates, as well as nonenzymatic functions, contributing to the full spectrum of events underlying their biological roles. We present two Kdm3a mouse models that exhibit cytoplasmic defects that may account in part for the globozoospermia phenotype reported previously. Electron microscopy revealed abnormal acrosome and manchette and the absence of implantation fossa at the caudal end of the nucleus in mice without Kdm3a demethylase activity, which affected cytoplasmic structures required to elongate the sperm head. We describe an enzymatically active new Kdm3a isoform and show that subcellular distribution, protein levels, and lysine demethylation activity of Kdm3a depended on Hsp90. We show that Kdm3a localizes to cytoplasmic structures of maturing spermatids affected in Kdm3a mutant mice, which in turn display altered fractionation of beta-actin and gamma-tubulin. Kdm3a is therefore a multifunctional Hsp90 client protein that participates directly in the regulation of cytoskeletal components.Publisher PDFPeer reviewe

Reorientation of Mispositioned Spindles in Short Astral Microtubule Mutant spc72Δ Is Dependent on Spindle Pole Body Outer Plaque and Kar3 Motor Protein

Nuclear migration and positioning in Saccharomyces cerevisiae depend on long astral microtubules emanating from the spindle pole bodies (SPBs). Herein, we show by in vivo fluorescence microscopy that cells lacking Spc72, the SPB receptor of the cytoplasmic γ-tubulin complex, can only generate very short (<1 μm) and unstable astral microtubules. Consequently, nuclear migration to the bud neck and orientation of the anaphase spindle along the mother-bud axis are absent in these cells. However, SPC72 deletion is not lethal because elongated but misaligned spindles can frequently reorient in mother cells, permitting delayed but otherwise correct nuclear segregation. High-resolution time-lapse sequences revealed that this spindle reorientation was most likely accomplished by cortex interactions of the very short astral microtubules. In addition, a set of double mutants suggested that reorientation was dependent on the SPB outer plaque and the astral microtubule motor function of Kar3 but not Kip2/Kip3/Dhc1, or the cortex components Kar9/Num1. Our observations suggest that Spc72 is required for astral microtubule formation at the SPB half-bridge and for stabilization of astral microtubules at the SPB outer plaque. In addition, our data exclude involvement of Spc72 in spindle formation and elongation functions

Supercurrents in Magnesium Diboride/Metal Composite Wire

We have fabricated a series of ex situ copper sheathed powder-in-tube MgB2 wires with 20% by volume Ag, Pb, In, and Ga metal added to the MgB2 powder. We find the transport critical current of these wires increases significantly with the addition of specific metals to the core filament. In particular, the critical current density (Jc) of the MgB2/Ga(20%) wire is in excess of 5x10^4 A/cm^2 at 10K in self field, nearly 50 times that of the MgB2/Ag(20%) wire. The temperature dependent Jc of all wires is well described as an ensemble of clean S/N/S junctions in which the relevant parameters are the average thickness of the N layer, the critical temperature of the S layer, and a scaling term related to Jc at zero temperature. Eliminating the differences in the filament microstructure as the primary cause of the enhanced Jc, we suggest that Jc is determined by the magnitude of the proximity effect induced superconductivity in the normal metal layer, which is known to be proportional to the electron-electron interaction in N. We present one-dimensional material specific calculations that support this, and zero-field cooled DC magnetic susceptibility data that confirm an increased number of well-connected superconducting grains exist in the composite wires that contain metal additions with large electron-electron interactions and long electron mean free paths.Comment: 36 pages, 10 figures, 3 table

Cnm67p Is a Spacer Protein of the Saccharomyces cerevisiae Spindle Pole Body Outer Plaque

In Saccharomyces cerevisiae, the spindle pole body (SPB) is the functional homolog of the mammalian centrosome, responsible for the organization of the tubulin cytoskeleton. Cytoplasmic (astral) microtubules essential for the proper segregation of the nucleus into the daughter cell are attached at the outer plaque on the SPB cytoplasmic face. Previously, it has been shown that Cnm67p is an integral component of this structure; cells deleted for CNM67 are lacking the SPB outer plaque and thus experience severe nuclear migration defects. With the use of partial deletion mutants of CNM67, we show that the N- and C-terminal domains of the protein are important for nuclear migration. The C terminus, not the N terminus, is essential for Cnm67p localization to the SPB. On the other hand, only the N terminus is subject to protein phosphorylation of a yet unknown function. Electron microscopy of SPB serial thin sections reveals that deletion of the N- or C-terminal domains disturbs outer plaque formation, whereas mutations in the central coiled-coil domain of Cnm67p change the distance between the SPB core and the outer plaque. We conclude that Cnm67p is the protein that connects the outer plaque to the central plaque embedded in the nuclear envelope, adjusting the space between them by the length of its coiled-coil