HSP16.6 Is Involved in the Development of Thermotolerance and Thylakoid Stability in the Unicellular Cyanobacterium, Synechocystis sp. PCC 6803

The low molecular weight (LMW) heat shock protein (HSP), HSP16.6, in the unicellular cyanobacterium, Synechocystis sp. PCC 6803, protects cells from elevated temperatures. A 95% reduction in the survival of mutant cells with an inactivated hsp16.6 was observed after exposure for 1 h at 47°C. Wild-type cell survival was reduced to only 41%. HSP16.6 is also involved in the development of thermotolerance. After a sublethal heat shock at 43°C for 1 h and subsequent challenge exposure at 49°C for 40 min, mutant cells did not survive, while 64% of wild-type cells survived. Ultrastructural changes in the integrity of thylakoid membranes of heat-shocked mutant cells also are discussed. These results demonstrate an important protective role for HSP16.6 in the protection of cells and, in particular, thylakoid membrane against thermal stress. Received: 14 October 1999 / Accepted: 16 November 1999     

Influence of polyploidy on insect herbivores of native and invasive genotypes of Solidago gigantea (Asteraceae)

Herbivores are sensitive to the genetic structure of plant populations, as genetics underlies plant phenotype and host quality. Polyploidy is a widespread feature of angiosperm genomes, yet few studies have examined how polyploidy influences herbivores. Introduction to new ranges, with consequent changes in selective regimes, can lead to evolution of changes in plant defensive characteristics and also affect herbivores. Here, we examine how insect herbivores respond to polyploidy in Solidago gigantea, using plants derived from both the native range (USA) and introduced range (Europe). S. gigantea has three cytotypes in the US, with two of these present in Europe. We performed bioassays with generalist (Spodoptera exigua) and specialist (Trirhabda virgata) leaf-feeding insects. Insects were reared on detached leaves (Spodoptera) or potted host plants (Trirhabda) and mortality and mass were measured. Trirhabda larvae showed little variation in survival or pupal mass attributable to either cytotype or plant origin. Spodoptera larvae were more sensitive to both cytotype and plant origin: they grew best on European tetraploids and poorly on US diploids (high mortality) and US tetraploids (low larval mass). These results show that both cytotype and plant origin influence insect herbivores, but that generalist and specialist insects may respond differently.Key words: polyploidy, cytotype, Solidago gigantea, insect herbivore, herbivory, invasive plant, introduced plantPolyploidy, or the possession of more than two sets of homologous chromosomes, is a fundamental force in angiosperm evolution.^1,2 Many plant species or species complexes consist of multiple cytotypes that may occur sympatrically;³ this is an important source of genetic structure in plant populations that is often overlooked.⁴ Possession of multiple genomes may confer advantages to polyploid plants such as increased heterozygosity, a decreased probability of inbreeding depression, or a greater gene pool available for selection; these traits contribute to the widespread success of polyploids and may make them prone to invasiveness.^5,6 In a recent article,⁷ we examined the functional consequences of polyploidy for different cytotypes of Solidago gigantea Ait. (Asteraceae), collected from both its native range (North America) and its introduced range (Europe). In this addendum, we show how cytotype and continent of origin influence interactions of S. gigantea with insect herbivores. Interactions with herbivores are expected to vary with cytotype because of phenotypic changes associated with polyploidy, but this area has received little study (reviewed in refs. ^8–11). Plant origin, from either the native range or an introduced range, should also influence herbivores. Plants may escape from their specialist natural enemies in the introduced range, thereby experiencing reduced herbivore pressure from an insect community dominated by generalists.^12,13 Given sufficient time, plants from the introduced range may evolve to decrease investment in anti-herbivore defenses, particularly those effective against specialists.¹⁴ While a growing body of research has addressed whether plant defenses against herbivory are lower in the introduced range,^12,15,16 few of these studies have also examined the influence of cytotype.¹⁷Three cytotypes of S. gigantea can be found in its native range in North America (diploid, tetraploid and hexaploid, 2n = 18, 36 and 54 respectively). These are morphologically indistinguishable and not generally treated as separate species.¹⁸ In Europe, where S. gigantea was introduced in the mid 18^th century,¹⁹ tetraploids are the dominant cytotype but diploids also occur. S. gigantea supports a diverse array of insect herbivores in its native range, but has few natural enemies in its introduced range.²⁰ We report here on experiments using both a generalist and a specialist leaf-chewing insect. The generalist, Spodoptera exigua (Lepidoptera: Noctuidae) is widely distributed and highly polyphagous, while the specialist Trirhabda virgata (Coleoptera: Chrysomelidae) feeds only on closely-related species within the genus Solidago. T. virgata is an outbreak insect that can be a major defoliator of S. gigantea and related species in North America.²¹ We grew plants originating from 10 populations in the US and 20 populations in Europe in common gardens at the University of Wisconsin-Milwaukee Field Station in Saukville, Wisconsin. There were five plant origin-cytotype combinations: three cytotypes from the US and two from Europe. Insects were reared on detached leaves from a single plant (Spodoptera) or on potted host plants (Trirhabda), for a set period of 21 d (Spodoptera) or until pupation (Trirhabda). We recorded insect survival and mass at the end of 21 d (Spodoptera) or at pupation (Trirhabda) (reviewed in ref. ²²).Overall survival was much better for the specialist Trirhabda than for the generalist Spodoptera (91% vs. 72%). Spodoptera larvae are not generally found on S. gigantea in the field, and while they are able to complete development, we found that this plant was not an ideal host. Spodoptera larvae were more sensitive to differences among cytotype and plant origin than were Trirhabda larvae. Percent survival was particularly poor for Spodoptera larvae reared on diploids from the US, where slightly more than half of the caterpillars survived for 21 days (Fig. 1). Trirhabda pupal mass was remarkably consistent across the five ploidy-plant origin combinations. In contrast, Spodoptera larvae responded to both cytotype and continent of origin. Surviving Spodoptera larvae did particularly well on tetraploid plants from the introduced range (Europe), and particularly poorly on tetraploids from the US (Fig. 1). We have previously reported that Spodoptera grow better on plants from Europe;²² our current results reveal that this difference is due exclusively to better growth on tetraploid plants. However, our results also show that both diploids and tetraploids from the US were poor hosts for Spodoptera: diploids because they caused high mortality and tetraploids because they resulted in poor growth. These results indicate that plants from the introduced range have reduced defenses against herbivores, even when accounting for polyploidy.Open in a separate windowFigure 1Mass ± se of S. exigua (A) and T. virgata (B) larvae reared on host plants of different cytotypes of Solidago gigantea originating from the US (native range) or europe (introduced range). Means in A followed by different letters are significantly different at p < 0.05 (ANOVA followed by multiple Student''s t-tests with Bonferroni correction). There were no significant differences in (B). Sample sizes for (A and B) shown in

CNS SIRT3 Expression Is Altered by Reactive Oxygen Species and in Alzheimer’s Disease

Progressive mitochondrial dysfunction contributes to neuronal degeneration in age-mediated disease. An essential regulator of mitochondrial function is the deacetylase, sirtuin 3 (SIRT3). Here we investigate a role for CNS Sirt3 in mitochondrial responses to reactive oxygen species (ROS)- and Alzheimer’s disease (AD)-mediated stress. Pharmacological augmentation of mitochondrial ROS increases Sirt3 expression in primary hippocampal culture with SIRT3 over-expression being neuroprotective. Furthermore, Sirt3 expression mirrors spatiotemporal deposition of β-amyloid in an AD mouse model and is also upregulated in AD patient temporal neocortex. Thus, our data suggest a role for SIRT3 in mechanisms sensing and tackling ROS- and AD-mediated mitochondrial stress.     

Peptidylarginine Deiminase 3 (PAD3) Is Upregulated by Prolactin Stimulation of CID-9 Cells and Expressed in the Lactating Mouse Mammary Gland

Peptidylarginine deiminases (PADs) post-translationally convert arginine into neutral citrulline residues. Our past work shows that PADs are expressed in the canine and murine mammary glands; however, the mechanisms regulating PAD expression and the function of citrullination in the normal mammary gland are unclear. Therefore, the first objective herein was to investigate regulation of PAD expression in mammary epithelial cells. We first examined PAD levels in CID-9 cells, which were derived from the mammary gland of mid-pregnant mice. PAD3 expression is significantly higher than all other PAD isoforms and mediates protein citrullination in CID-9 cells. We next hypothesized that prolactin regulates PAD3 expression. To test this, CID-9 cells were stimulated with 5 μg/mL of prolactin for 48 hours which significantly increases PAD3 mRNA and protein expression. Use of a JAK2 inhibitor and a dominant negative (DN)-STAT5 adenovirus indicate that prolactin stimulation of PAD3 expression is mediated by the JAK2/STAT5 signaling pathway in CID-9 cells. In addition, the human PAD3 gene promoter is prolactin responsive in CID-9 cells. Our second objective was to investigate the expression and activity of PAD3 in the lactating mouse mammary gland. PAD3 expression in the mammary gland is highest on lactation day 9 and coincident with citrullinated proteins such as histones. Use of the PAD3 specific inhibitor, Cl4-amidine, indicates that PAD3, in part, can citrullinate proteins in L9 mammary glands. Collectively, our results show that upregulation of PAD3 is mediated by prolactin induction of the JAK2/STAT5 signaling pathway, and that PAD3 appears to citrullinate proteins during lactation.     

A mechanism of expansion: Arctic deciduous shrubs capitalize on warming-induced nutrient availability

Warming-induced nutrient enrichment in the Arctic may lead to shifts in leaf-level physiological properties and processes with potential consequences for plant community dynamics and ecosystem function. To explore the physiological responses of Arctic tundra vegetation to increasing nutrient availability, we examined how a set of leaf nutrient and physiological characteristics of eight plant species (representing four plant functional groups) respond to a gradient of experimental nitrogen (N) and phosphorus (P) enrichment. Specifically, we examined a set of chlorophyll fluorescence measures related to photosynthetic efficiency, performance and stress, and two leaf nutrient traits (leaf %C and %N), across an experimental nutrient gradient at the Arctic Long Term Ecological Research site, located in the northern foothills of the Brooks Range, Alaska. In addition, we explicitly assessed the direct relationships between chlorophyll fluorescence and leaf %N. We found significant differences in physiological and nutrient traits between species and plant functional groups, and we found that species within one functional group (deciduous shrubs) have significantly greater leaf %N at high levels of nutrient addition. In addition, we found positive, saturating relationships between leaf %N and chlorophyll fluorescence measures across all species. Our results highlight species-specific differences in leaf nutrient traits and physiology in this ecosystem. In particular, the effects of a gradient of nutrient enrichment were most prominent in deciduous plant species, the plant functional group known to be increasing in relative abundance with warming in this ecosystem.     

Genetically Engineered Mesenchymal Stem Cells as a Proposed Therapeutic for Huntington’s Disease

There is much interest in the use of mesenchymal stem cells/marrow stromal cells (MSC) to treat neurodegenerative disorders, in particular those that are fatal and difficult to treat, such as Huntington's disease. MSC present a promising tool for cell therapy and are currently being tested in FDA-approved phase I-III clinical trials for many disorders. In preclinical studies of neurodegenerative disorders, MSC have demonstrated efficacy, when used as delivery vehicles for neural growth factors. A number of investigators have examined the potential benefits of innate MSC-secreted trophic support and augmented growth factors to support injured neurons. These include overexpression of brain-derived neurotrophic factor and glial-derived neurotrophic factor, using genetically engineered MSC as a vehicle to deliver the cytokines directly into the microenvironment. Proposed regenerative approaches to neurological diseases using MSC include cell therapies in which cells are delivered via intracerebral or intrathecal injection. Upon transplantation, MSC in the brain promote endogenous neuronal growth, encourage synaptic connection from damaged neurons, decrease apoptosis, reduce levels of free radicals, and regulate inflammation. These abilities are primarily modulated through paracrine actions. Clinical trials for MSC injection into the central nervous system to treat amyotrophic lateral sclerosis, traumatic brain injury, and stroke are currently ongoing. The current data in support of applying MSC-based cellular therapies to the treatment of Huntington's disease is discussed.     

Genome-Wide Association Studies in an Isolated Founder Population from the Pacific Island of Kosrae

It has been argued that the limited genetic diversity and reduced allelic heterogeneity observed in isolated founder populations facilitates discovery of loci contributing to both Mendelian and complex disease. A strong founder effect, severe isolation, and substantial inbreeding have dramatically reduced genetic diversity in natives from the island of Kosrae, Federated States of Micronesia, who exhibit a high prevalence of obesity and other metabolic disorders. We hypothesized that genetic drift and possibly natural selection on Kosrae might have increased the frequency of previously rare genetic variants with relatively large effects, making these alleles readily detectable in genome-wide association analysis. However, mapping in large, inbred cohorts introduces analytic challenges, as extensive relatedness between subjects violates the assumptions of independence upon which traditional association test statistics are based. We performed genome-wide association analysis for 15 quantitative traits in 2,906 members of the Kosrae population, using novel approaches to manage the extreme relatedness in the sample. As positive controls, we observe association to known loci for plasma cholesterol, triglycerides, and C-reactive protein and to a compelling candidate loci for thyroid stimulating hormone and fasting plasma glucose. We show that our study is well powered to detect common alleles explaining ≥5% phenotypic variance. However, no such large effects were observed with genome-wide significance, arguing that even in such a severely inbred population, common alleles typically have modest effects. Finally, we show that a majority of common variants discovered in Caucasians have indistinguishable effect sizes on Kosrae, despite the major differences in population genetics and environment.