Identification of amino acid residues of mammalian mitochondrial phosphate carrier important for its functional expression in yeast cells, as achieved by PCR-mediated random mutation and gap-repair cloning

The mitochondrial phosphate carrier (PiC) of mammals, but not the yeast one, is synthesized with a presequence. The deletion of this presequence of the mammalian PiC was reported to facilitate the import of the carrier into yeast mitochondria, but the question as to whether or not mammalian PiC could be functionally expressed in yeast mitochondria was not addressed. In the present study, we first examined whether the defective growth on a glycerol plate of yeast cells lacking the yeast PiC gene could be reversed by the introduction of expression vectors of rat PiCs. The introduction of expression vectors encoding full-length rat PiC (rPiC) or rPiC lacking the presequence (ΔNrPiC) was ineffective in restoring growth on the glycerol plates. When we examined the expression levels of individual rPiCs in yeast mitochondria, ΔNrPiC was expressed at a level similar to that of yeast PiC, but that of rPiC was very low. These results indicated that ΔNrPiC expressed in yeast mitochondria is inert. Next, we sought to isolate “revertants” viable on the glycerol plate by expressing randomly mutated ΔNrPiC, and obtained two clones. These clones carried either of two mutations, F267S or F282S; and these mutations restored the transport function of ΔNrPiC in yeast mitochondria. These two Phe residues were conserved in human carrier (hPiC), and the transport function of ΔNhPiC expressed in yeast mitochondria was also markedly improved by their substitutions. Thus, substitution of F267S or F282S was concluded to be important for functional expression of mammalian PiCs in yeast mitochondria

Insect Adenine Nucleotide Translocases

Mitochondrial adenine nucleotide translocase (ANT) specifically acts in ADP/ATP exchange through the mitochondrial inner membrane. This transporter protein thereby plays a significant role in energy metabolism in eukaryotic cells. Most mammals have four paralogous ANT genes (ANT1-4) and utilize these paralogues in different types of cells. The fourth paralogue of ANT (ANT4) is present only in mammals and reptiles and is exclusively expressed in testicular germ cells where it is required for meiotic progression in the spermatocytes. Here, we report that silkworms harbor two ANT paralogues, the homeostatic paralogue (BmANTI1) and the testis-specific paralogue (BmANTI2). The BmANTI2 protein has an N-terminal extension in which the positions of lysine residues in the amino acid sequence are distributed as in human ANT4. An expression analysis showed that BmANTI2 transcripts were restricted to the testis, suggesting the protein has a role in the progression of spermatogenesis. By contrast, BmANTI1 was expressed in all tissues tested, suggesting it has an important role in homeostasis. We also observed that cultured silkworm cells required BmANTI1 for proliferation. The ANTI1 protein of the lepidopteran Plutella xylostella (PxANTI1), but not those of other insect species (or PxANTI2), restored cell proliferation in BmANTI1-knockdown cells suggesting that ANTI1 has similar energy metabolism functions across the Lepidoptera. Our results suggest that BmANTI2 is evolutionarily divergent from BmANTI1 and has developed a specific role in spermatogenesis similar to that of mammalian ANT4

Silverrush. Xii. Intensity Mapping for Ly Α Emission Extending over 100-1000 Comoving Kpc Around Z ∼2-7 Laes with Subaru Hsc-Ssp and Chorus Data

We conduct intensity mapping to probe for extended diffuse Lyα emission around Lyα emitters (LAEs) at z ∼2-7, exploiting very deep (∼26 mag at 5σ) and large-area (∼4.5 deg2) Subaru/Hyper Suprime-Cam narrowband (NB) images and large LAE catalogs consisting of a total of 1540 LAEs at z = 2.2, 3.3, 5.7, and 6.6 obtained by the HSC-SSP and CHORUS projects. We calculate the spatial correlations of these LAEs with ∼1-2-billion-pixel flux values of the NB images, deriving the average Lyα surface brightness (SBLyα ) radial profiles around the LAEs. By carefully estimating systematics such as fluctuations of sky background and point-spread functions, we detect Lyα emission at 100-1000 comoving kpc around z = 3.3 and 5.7 LAEs at the 3.2σ and 3.7σ levels, respectively, and tentatively (=2.0σ) at z = 6.6. The emission is as diffuse as ∼10-20-10-19 erg s-1 cm-2 arcsec-2 and extended beyond the virial radius of a dark matter halo with a mass of 1011 M. While the observed SBLyα profiles have similar amplitudes at z = 2.2-6.6 within the uncertainties, the intrinsic SBLyα profiles (corrected for the cosmological dimming effect) increase toward high redshifts. This trend may be explained by increasing hydrogen gas density due to the evolution of the cosmic volume. Comparisons with theoretical models suggest that extended Lyα emission around an LAE is powered by resonantly scattered Lyα photons in the CGM and IGM that originate from the inner part of the LAE and/or neighboring galaxies around the LAE

Differential contribution of the Fanconi anemia-related proteins to repair of several types of DNA damage in cultured silkworm cells

AbstractThe silkworm Fanconi anemia (FA) pathway is required for normal cellular resistance to mitomycin C (MMC) in silkworms, but little is known about the requirement for repair of other types of DNA damage. Here we report that silkworm cells deficient for FA proteins FancD2 and FancM exhibit normal sensitivities to hydroxyurea (HU) and camptothecin (CPT), although FancM-dependent FancD2 monoubiquitination is induced upon these treatments. Similar results were observed in cells depleted for Rmi1 and Mhf1, which interact with the FancM protein. We also found that Rad51-knockdown cells exhibited normal sensitivity to HU despite induction of double-strand breaks by HU treatment

Kinematic Design and Analysis of a Wearable End-Effector Type Upper Limb Assistive Robot

A novel wearable end-effector type upper limb assistive robot for stroke hemiplegic patients is proposed, and a 3 DOF 4R-5R parallel mechanism for this type of robots is presented. Assistive functions of the robot are finalized to motion and force assistance to allow patients to have their paralyzed limb perform auxiliary roles in ADL. Kinematics of this mechanism, including joint configuration and placement of three active joints, is presented, followed by displacement analysis and workspace analysis. Motion of the proposed mechanism is confirmed by an experimental prototype

Two Adenine Nucleotide Translocase Paralogues Involved in Cell Proliferation and Spermatogenesis in the Silkworm <i>Bombyx mori</i>

<div><p>Mitochondrial adenine nucleotide translocase (ANT) specifically acts in ADP/ATP exchange through the mitochondrial inner membrane. This transporter protein thereby plays a significant role in energy metabolism in eukaryotic cells. Most mammals have four paralogous <i>ANT</i> genes (<i>ANT1-4</i>) and utilize these paralogues in different types of cells. The fourth paralogue of <i>ANT</i> (<i>ANT4</i>) is present only in mammals and reptiles and is exclusively expressed in testicular germ cells where it is required for meiotic progression in the spermatocytes. Here, we report that silkworms harbor two ANT paralogues, the homeostatic paralogue (BmANTI1) and the testis-specific paralogue (BmANTI2). The BmANTI2 protein has an N-terminal extension in which the positions of lysine residues in the amino acid sequence are distributed as in human ANT4. An expression analysis showed that BmANTI2 transcripts were restricted to the testis, suggesting the protein has a role in the progression of spermatogenesis. By contrast, BmANTI1 was expressed in all tissues tested, suggesting it has an important role in homeostasis. We also observed that cultured silkworm cells required BmANTI1 for proliferation. The ANTI1 protein of the lepidopteran <i>Plutella xylostella</i> (PxANTI1), but not those of other insect species (or PxANTI2), restored cell proliferation in BmANTI1-knockdown cells suggesting that ANTI1 has similar energy metabolism functions across the Lepidoptera. Our results suggest that BmANTI2 is evolutionarily divergent from BmANTI1 and has developed a specific role in spermatogenesis similar to that of mammalian ANT4.</p></div