Potassium/proton antiport system of Escherichia coli

The intracellular level of potassium (K(+)) in Escherichia coli is regulated through multiple K(+) transport systems. Recent data indicate that not all K(+) extrusion system(s) have been identified (15). Here we report that the E. coli Na(+) (Ca(2+))/H(+) antiporter ChaA functions as a K(+) extrusion system. Cells expressing ChaA mediated K(+) efflux against a K(+) concentration gradient. E. coli strains lacking the chaA gene were unable to extrude K(+) under conditions in which wild-type cells extruded K(+). The K(+)/H(+) antiporter activity of ChaA was detected by using inverted membrane vesicles produced using a French press. Physiological growth studies indicated that E. coli uses ChaA to discard excessive K(+), which is toxic for these cells. These results suggest that ChaA K(+)/H(+) antiporter activity enables E. coli to adapt to K(+) salinity stress and to maintain K(+) homeostasis.     

Single gene deletions of mrpA to mrpG and mrpE point mutations affect activity of the Mrp Na+/H+ antiporter of alkaliphilic Bacillus and formation of hetero-oligomeric Mrp complexes

Mrp antiporters catalyze secondary Na(+)(Li(+))/H(+) antiport and/or K(+)/H(+) antiport that is physiologically important in diverse bacteria. An additional capacity for anion flux has been observed for a few systems. Mrp is unique among antiporters in that it requires all six or seven hydrophobic gene products (MrpA to MrpG) of the mrp operon for full antiporter activity, but MrpE has been reported to be dispensable. Here, the membrane complexes formed by Mrp proteins were examined using a cloned mrp operon from alkaliphilic Bacillus pseudofirmus OF4. The operon was engineered so that the seven Mrp proteins could be detected in single samples. Membrane extracts of an antiporter-deficient Escherichia coli strain expressing this construct were analyzed by blue native-sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Mrp complexes of two sizes were identified containing all seven Mrp proteins. Studies of the single nonpolar mrp gene deletions in the construct showed that a subcomplex of MrpA, MrpB, MrpC, and MrpD was formed in the absence of MrpE, MrpF, or MrpG. By contrast, MrpE, MrpF, and MrpG were not observed in membranes lacking MrpA, MrpB, MrpC, or MrpD. Although MrpA and MrpD have been hypothesized to be the antiporter proteins, the MrpA-to-D complex was inactive. Every Mrp protein was required for an activity level near that of the wild-type Na(+)/H(+) antiporter, but a very low activity level was observed in the absence of MrpE. The introduction of an MrpE(P114G) mutation into the full Mrp complex led to antiport activity with a greatly increased apparent K(m) value for Na(+). The results suggested that interactions among the proteins of heterooligomeric Mrp complexes strongly impact antiporter properties.     

Involvement of IL-1 in the Maintenance of Masseter Muscle Activity and Glucose Homeostasis

Physical exercise reportedly stimulates IL-1 production within working skeletal muscles, but its physiological significance remains unknown due to the existence of two distinct IL-1 isoforms, IL-1α and IL-1β. The regulatory complexities of these two isoforms, in terms of which cells in muscles produce them and their distinct/redundant biological actions, have yet to be elucidated. Taking advantage of our masticatory behavior (Restrained/Gnawing) model, we herein show that IL-1α/1β-double-knockout (IL-1-KO) mice exhibit compromised masseter muscle (MM) activity which is at least partially attributable to abnormalities of glucose handling (rapid glycogen depletion along with impaired glucose uptake) and dysfunction of IL-6 upregulation in working MMs. In wild-type mice, masticatory behavior clearly increased IL-1β mRNA expression but no incremental protein abundance was detectable in whole MM homogenates, whereas immunohistochemical staining analysis revealed that both IL-1α- and IL-1β-immunopositive cells were recruited around blood vessels in the perimysium of MMs after masticatory behavior. In addition to the aforementioned phenotype of IL-1-KO mice, we found the IL-6 mRNA and protein levels in MMs after masticatory behavior to be significantly lower in IL-1-KO than in WT. Thus, our findings confirm that the locally-increased IL-1 elicited by masticatory behavior, although present small in amounts, contributes to supporting MM activity by maintaining normal glucose homeostasis in these muscles. Our data also underscore the importance of IL-1-mediated local interplay between autocrine myokines including IL-6 and paracrine cytokines in active skeletal muscles. This interplay is directly involved in MM performance and fatigability, perhaps mediated through maintaining muscular glucose homeostasis.     

High Prevalence of Simian T-Lymphotropic Virus Type L in Wild Ethiopian Baboons

Simian T-cell leukemia viruses (STLVs) are the simian counterparts of human T-cell leukemia viruses (HTLVs). A novel, divergent type of STLV (STLV-L) from captive baboons was reported in 1994, but its natural prevalence remained unclear. We investigated the prevalence of STLV-L in 519 blood samples from wild-living nonhuman primates in Ethiopia. Seropositive monkeys having cross-reactive antibodies against HTLV were found among 22 out of 40 hamadryas baboons, 8 of 96 anubis baboons, 24 of 50 baboons that are hybrids between hamadryas and anubis baboons, and 41 of 177 grivet monkeys, but not in 156 gelada baboons. A Western blotting assay showed that sera obtained from seropositive hamadryas and hybrid baboons exhibited STLV-L-like reactivity. A PCR assay successfully amplified STLV sequences, which were subsequently sequenced and confirmed as being closely related to STLV-L. Surprisingly, further PCR showed that nearly half of the hamadryas (20 out of 40) and hybrid (19 out of 50) baboons had STLV-L DNA sequences. In contrast, most of the seropositive anubis baboons and grivet monkeys carried typical STLV-1 but not STLV-L. These observations demonstrate that STLV-L naturally prevails among hamadryas and hybrid baboons at significantly high rates. STLV-1 and -2, the close relative of STLV-L, are believed to have jumped across simian-human barriers, which resulted in widespread infection of HTLV-1 and -2. Further studies are required to know if STLV-L is spreading into human populations.     

Diverse variation of reproductive barriers in three intraspecific rice crosses

Reproductive barriers are thought to play an important role in the processes of speciation and differentiation. Asian rice cultivars, Oryza sativa, can be classified into two main types, Japonica and Indica, on the basis of several characteristics. The fertility of Japonica-Indica hybrids differs from one cross to another. Many genes involved in reproductive barriers (hybrid sterility, hybrid weakness, and gametophytic competition genes) have been reported in different Japonica-Indica crosses. To clarify the state of Japonica-Indica differentiation, all reproductive barriers causing deviation from Mendelian segregation ratios in F(2) populations were mapped and compared among three different Japonica-Indica crosses: Nipponbare/Kasalath (NK), Fl1084/Dao Ren Qiao (FD), and Fl1007/Kinandang puti (FK). Mapping of reproductive barriers was performed by regression analysis of allele frequencies of DNA markers covering the entire genome. Allele frequencies were explained by 33 reproductive barriers (15 gametophytic and 18 zygotic) in NK, 32 barriers (15 gametophytic and 17 zygotic) in FD, and 37 barriers (19 gametophytic and 18 zygotic) in FK. The number of reproductive barriers in the three crosses was similar; however, most of the barriers were mapped at different loci. Therefore, these reproductive barriers formed after Japonica-Indica differentiation. Considering the high genetic similarity within Japonica and Indica cultivars, the differences in the reproductive barriers of each cross were unexpectedly numerous. The reproductive barriers of Japonica-Indica hybrids likely evolved more rapidly than other genetic elements. One possible force responsible for such rapid evolution of the barriers may have been the domestication of rice.     

Intraspecific variation in the status of ant symbiosis on a myrmecophyte,<Emphasis Type="Italic"> Macaranga bancana,</Emphasis> between primary and secondary forests in Borneo

A tree species, Macaranga bancana , distributed in South East Asian tropics has a mutualistic relationship with specific symbiotic ant species, which defend the plant from herbivores. To examine the intraspecific variation in the status of the ant-plant symbiosis among microhabitats of different light conditions, we investigated the species composition of nesting ants and the herbivory damage on M. bancana saplings by field observations and sampling in primary and secondary forests in Sarawak. In addition, the effectiveness of non-ant (physical and chemical) defenses were estimated by feeding the larvae of a polyphagous lepidopteran with M. bancana leaves from saplings in the two types of forests. All saplings in the primary forest were colonized by two Crematogaster ant species that had been known to be the obligate symbionts of M. bancana, while in the secondary forest, about half of the saplings were occupied by several ant species that were not obligate symbionts. There was little herbivory damage on saplings colonized by the two Crematogaster symbiont ants in both forest types, while the saplings colonized by the other ant species suffered a 10–60% loss of leaf area. Larval mortality of the polyphagous lepidopteran Spodoptera litura was significantly higher when larvae fed on leaves of M. bancana saplings in the secondary forest than when fed on leaves of M. bancana saplings in the primary forest. These results suggest that the symbiosis between ants and M. bancana is looser and the non-ant-defenses are stronger in secondary forests, where light is more intense, than in primary forests.     

Three-dimensional solution structure of an archaeal FKBP with a dual function of peptidyl prolyl cis-trans isomerase and chaperone-like activities

Here we report the solution structure of an archaeal FK506-binding protein (FKBP) from a thermophilic archaeum, Methanococcus thermolithotrophicus (MtFKBP17), which has peptidyl prolyl cis-trans isomerase (PPIase) and chaperone-like activities, to reveal the structural basis for the dual function. In addition to a typical PPIase domain, a newly identified domain is formed in the flap loop by a 48-residue insert that is required for the chaperone-like activity. The new domain, called IF domain (the Insert in the Flap), is a novel-folding motif and exposes a hydrophobic surface, which we consider to play an important role in the chaperone-like activity.