Two new endemic species of Ophiorrhiza L. (Rubiaceae: Ophiorrhizeae) from Davao Oriental,Philippines

Two new species of Ophiorrhiza collected from Mt Hamiguitan Range Wildlife Sanctuary in Davao Oriental, Philippines, are herein described and illustrated. Ophiorrhiza erythropilosa is the third Philippine species possessing involucral bracts and is further characterized by its predominantly villose stems that appear red due to strikingly red-violet trichomes, subpersistent bifid stipules, linear involucral bracts without prominent midrib, and 4 mm long urceolate corolla that is villose outside and lightly puberulous inside. Ophiorrhiza hamiguitanensis is characterized by its coriaceous, lanceolate leaves with attenuate apex and base, brochidodromous venation, subpersistent bilobed stipules with slightly recurved acute tips, presence of 1.5–2.5 mm long linear ensiform bracts, heterostylous flowers, clavate calyces and broadly obcordate capsules.     

Animal movements in fire‐prone landscapes

Movement is a trait of fundamental importance in ecosystems subject to frequent disturbances, such as fire‐prone ecosystems. Despite this, the role of movement in facilitating responses to fire has received little attention. Herein, we consider how animal movement interacts with fire history to shape species distributions. We consider how fire affects movement between habitat patches of differing fire histories that occur across a range of spatial and temporal scales, from daily foraging bouts to infrequent dispersal events, and annual migrations. We review animal movements in response to the immediate and abrupt impacts of fire, and the longer‐term successional changes that fires set in train. We discuss how the novel threats of altered fire regimes, landscape fragmentation, and invasive species result in suboptimal movements that drive populations downwards. We then outline the types of data needed to study animal movements in relation to fire and novel threats, to hasten the integration of movement ecology and fire ecology. We conclude by outlining a research agenda for the integration of movement ecology and fire ecology by identifying key research questions that emerge from our synthesis of animal movements in fire‐prone ecosystems.     

Yeast Fex1p Is a Constitutively Expressed Fluoride Channel with Functional Asymmetry of Its Two Homologous Domains

Fluoride is a ubiquitous environmental toxin with which all biological species must cope. A recently discovered family of fluoride export (FEX) proteins protects organisms from fluoride toxicity by removing it from the cell. We show here that FEX proteins in Saccharomyces cerevisiae function as ion channels that are selective for fluoride over chloride and that these proteins are constitutively expressed at the yeast plasma membrane. Continuous expression is in contrast to many other toxin exporters in yeast, and this, along with the fact that two nearly duplicate proteins are encoded in the yeast genome, suggests that the threat posed by fluoride ions is frequent and detrimental. Structurally, eukaryotic FEX proteins consist of two homologous four-transmembrane helix domains folded into an antiparallel dimer, where the orientation of the two domains is fixed by a single transmembrane linker helix. Using phylogenetic sequence conservation as a guide, we have identified several functionally important residues. There is substantial functional asymmetry in the effect of mutation at corresponding sites in the two domains. Specifically, mutations to residues in the C-terminal domain proved significantly more detrimental to function than did similar mutations in the N-terminal domain. Our data suggest particular residues that may be important to anion specificity, most notably the necessity of a positive charge near the end of TMH1 in the C-terminal domain. It is possible that a cationic charge at this location may create an electrostatic well for fluoride ions entering the channel from the cytoplasm.     

Mutation in HSF4 associated with early but not late-onset hereditary cataract in the Boston Terrier

Primary hereditary cataract (HC) is one of the most common disorders in purebred dogs and is a leading cause of blindness. Boston Terriers suffer from 2 distinct forms of HC which occur at different ages and which are different in their appearance and progression. Early-onset hereditary cataract (EHC) affects dogs within the first few months of life, is always progressive and bilateral, and results in total blindness, whereas late-onset hereditary cataract (LHC) in general affects dogs over the age of 3 and is more variable in its clinical phenotype, age of onset, progression, and the degree to which vision is impaired. A mutation in HSF4 has recently been reported in a small number of Boston Terriers affected with EHC. In this study, we analyzed 22 additional Boston Terriers affected with early-onset cataract to confirm that the HSF4 mutation is causative for this form of cataract in this breed. In addition, we analyzed 40 Boston Terriers that were either clinically clear or affected with LHC for the presence or absence of the HSF4 mutation. By also sequencing HSF4 in dogs affected with LHC, we conclude that HSF4 is not associated with the development of the late-onset form of cataract and that the 2 forms of cataract in this breed are therefore genetically discrete conditions.     

Identification and characterization of a suppressor T cell hybridoma specifically inducible by L-glutamic acid60-L-alanine30-L-tyrosine10 (GAT)

In vitro activation of naive spleen cells from C57BL/10 mice with GAT and the monoclonal GAT-TsF1, 372B3.5, followed by fusion with BW5147 resulted in generation of a hybridoma that fails to produce GAT-TsF constitutively, but upon reexposure to GAT and 372B3.5 is induced to secrete GAT-TsF2. The induction is GAT specific and requires de novo RNA, protein synthesis, and DNA synthesis. Although both GAT and 372B3.5 are required for induction, they may be added sequentially, provided the GAT is added first. The GAT-TsF produced by the induced cell is antigen specific and composed of two polypeptide chains: one capable of binding antigen, the other bearing determinants encoded by the I-J region of the MHC. The utility of this inducible GAT-TsF2 cell line for molecular biology and other studies is discussed.     

ESCRT-I Function is Required for Tyrp1 Transport from Early Endosomes to the Melanosome Limiting Membrane

Melanosomes are lysosome-related organelles that coexist with lysosomes within melanocytes. The pathways by which melanosomal proteins are diverted from endocytic organelles toward melanosomes are incompletely defined. In melanocytes from mouse models of Hermansky-Pudlak syndrome that lack BLOC-1, melanosomal proteins such as tyrosinase-related protein 1 (Tyrp1) accumulate in early endosomes. Whether this accumulation represents an anomalous pathway or an arrested normal intermediate in melanosome protein trafficking is not clear. Here, we show that early endosomes are requisite intermediates in the trafficking of Tyrp1 from the Golgi to late stage melanosomes in normal melanocytic cells. Kinetic analyses show that very little newly synthesized Tyrp1 traverses the cell surface and that internalized Tyrp1 is inefficiently sorted to melanosomes. Nevertheless, nearly all Tyrp1 traverse early endosomes since it becomes trapped within enlarged, modified endosomes upon overexpression of Hrs. Although Tyrp1 localization is not affected by Hrs depletion, depletion of the ESCRT-I component, Tsg101, or inhibition of ESCRT function by dominant-negative approaches results in a dramatic redistribution of Tyrp1 to aberrant endosomal membranes that are largely distinct from those harboring traditional ESCRT-dependent, ubiquitylated cargoes such as MART-1. The lysosomal protein content of some of these membranes and the lack of Tyrp1 recycling to the plasma membrane in Tsg101-depleted cells suggests that ESCRT-I functions downstream of BLOC-1. Our data delineate a novel pathway for Tyrp1 trafficking and illustrate a requirement for ESCRT-I function in controlling protein sorting from vacuolar endosomes to the limiting membrane of a lysosome-related organelle.     

Antibody Profiling by Luciferase Immunoprecipitation Systems (LIPS)

Technologies for comprehensively understanding and quantifying antibody profiles to autoantigens and infectious agents may yield new insights into disease mechanisms and may elucidate new markers to substratify disease with different clinical features and better understand pathogenesis. We have developed a highly quantitative method called Luciferase Immunoprecipitation Systems (LIPS) for profiling patient sera antibody responses to autoantigens and pathogen antigens associated with infection. Unlike ELISAs, the highly sensitive LIPS is easily implemented to survey humoral serological response profiles to different antigens in a universal format and produces dynamic antibody titer ranges up to 6 log₁₀ for some antigens. In these studies, quantitative profiling by LIPS of patient humoral responses against panels of antigens or even the entire proteome of some pathogens (i.e. HIV), is typically more informative than testing a single antigen by ELISA. In addition, LIPS also eliminates time and effort needed to produce highly purified antigens as well as the labor-intensive assay optimization steps needed for standard ELISAs. Here we provide a detailed protocol describing the technical aspects of performing LIPS assays for readily profiling antibody responses to single or multiple antigens.Download video file.^{(144M, mp4)}