Genetics and evolution of the chloroplast : (cytoplasmic inheritance, nucleus-cytoplasm interaction, plastome recombination, plastome mutator, oenothera)

As a photosynthetic organelle and as the site of several important biosynthetic pathways, the chloroplast is essential to the survival of the plant cell. In evolutionary terms, the chloroplast genetic material can be considered conservative: from the green algae to higher plants, chloroplast DNAs have similar sizes, gene contents and arrangements, although some modifications of the chloroplast DNA have occurred as it has coevolved with the nucleus. Mutation and recombination are traditionally considered responsible for evolutionary change, but these may be quite limited in the chloroplast genetic system. Gene conversion may result in the elimination of new mutations, while the widespread occurrence of uniparental inheritance of chloroplasts reduces the likelihood of two different plastids meeting and their DNAs recombining. In other systems, transposons contribute to genetic change. One possible explanation for plastid mutator genes is that they activate transposable elements in the chloroplast DNA. Among diverse genera in primitive and higher plants, both uniparental and biparental transmission of chloroplasts occur in sexual crosses. When maternal inheritance of plastids occurs, the paternal plastids may be excluded or eliminated from the male gamete in a variety of ways. Thus maternal inheritance has probably appeared in the algae, mosses, ferns, gymnosperms, and angiosperms through parallel evolution.BARBARA B. SEARS, Department of Botany and Plant Pathology, Michigan State University, East Lansing, Michigan

Chloroplast lipid transfer processes in \u3ci\u3eChlamydomonas reinhardtii\u3c/i\u3e involving a TRIGALACTOSYLDIACYLGLYCEROL 2 (TGD2) ortholog

In plants, lipids of the photosynthetic membrane are synthesized by parallel pathways associated with the endoplasmic reticulum (ER) and the chloroplast envelope membranes. Lipids derived from the two pathways are distinguished by their acyl-constituents. Following this plant paradigm, the prevalent acyl composition of chloroplast lipids suggests that Chlamydomonas reinhardtii (Chlamydomonas) does not use the ER pathway; however, the Chlamydomonas genome encodes presumed plant orthologs of a chloroplast lipid transporter consisting of TGD (TRIGALACTOSYLDIACYLGLYCEROL) proteins that are required for ER-tochloroplast lipid trafficking in plants. To resolve this conundrum, we identified a mutant of Chlamydomonas deleted in the TGD2 gene and characterized the respective protein, CrTGD2. Notably, the viability of the mutant was reduced, showing the importance of CrTGD2. Galactoglycerolipid metabolism was altered in the tgd2 mutant with monogalactosyldiacylglycerol (MGDG) synthase activity being strongly stimulated. We hypothesize this to be a result of phosphatidic acid accumulation in the chloroplast outer envelope membrane, the location of MGDG synthase in Chlamydomonas. Concomitantly, increased conversion of MGDG into triacylglycerol (TAG) was observed. This TAG accumulated in lipid droplets in the tgd2 mutant under normal growth conditions. Labeling kinetics indicate that Chlamydomonas can import lipid precursors from the ER, a process that is impaired in the tgd2 mutant. Includes Supplementary Material

Genome, Functional Gene Annotation, and Nuclear Transformation of the Heterokont Oleaginous Alga \u3ci\u3eNannochloropsis oceanica\u3c/i\u3e CCMP1779

Unicellular marine algae have promise for providing sustainable and scalable biofuel feedstocks, although no single species has emerged as a preferred organism. Moreover, adequate molecular and genetic resources prerequisite for the rational engineering of marine algal feedstocks are lacking for most candidate species. Heterokonts of the genus Nannochloropsis naturally have high cellular oil content and are already in use for industrial production of high-value lipid products. First success in applying reverse genetics by targeted gene replacement makes Nannochloropsis oceanica an attractive model to investigate the cell and molecular biology and biochemistry of this fascinating organism group. Here we present the assembly of the 28.7 Mb genome of N. oceanica CCMP1779. RNA sequencing data from nitrogen-replete and nitrogendepleted growth conditions support a total of 11,973 genes, of which in addition to automatic annotation some were manually inspected to predict the biochemical repertoire for this organism. Among others, more than 100 genes putatively related to lipid metabolism, 114 predicted transcription factors, and 109 transcriptional regulators were annotated. Comparison of the N. oceanica CCMP1779 gene repertoire with the recently published N. gaditana genome identified 2,649 genes likely specific to N. oceanica CCMP1779. Many of these N. oceanica–specific genes have putative orthologs in other species or are supported by transcriptional evidence. However, because similarity-based annotations are limited, functions of most of these species-specific genes remain unknown. Aside from the genome sequence and its analysis, protocols for the transformation of N. oceanica CCMP1779 are provided. The availability of genomic and transcriptomic data for Nannochloropsis oceanica CCMP1779, along with efficient transformation protocols, provides a blueprint for future detailed gene functional analysis and genetic engineering of Nannochloropsis species by a growing academic community focused on this genus

Primary brain calcification: an international study reporting novel variants and associated phenotypes.

Primary familial brain calcification (PFBC) is a rare cerebral microvascular calcifying disorder with a wide spectrum of motor, cognitive, and neuropsychiatric symptoms. It is typically inherited as an autosomal-dominant trait with four causative genes identified so far: SLC20A2, PDGFRB, PDGFB, and XPR1. Our study aimed at screening the coding regions of these genes in a series of 177 unrelated probands that fulfilled the diagnostic criteria for primary brain calcification regardless of their family history. Sequence variants were classified as pathogenic, likely pathogenic, or of uncertain significance (VUS), based on the ACMG-AMP recommendations. We identified 45 probands (25.4%) carrying either pathogenic or likely pathogenic variants (n = 34, 19.2%) or VUS (n = 11, 6.2%). SLC20A2 provided the highest contribution (16.9%), followed by XPR1 and PDGFB (3.4% each), and PDGFRB (1.7%). A total of 81.5% of carriers were symptomatic and the most recurrent symptoms were parkinsonism, cognitive impairment, and psychiatric disturbances (52.3%, 40.9%, and 38.6% of symptomatic individuals, respectively), with a wide range of age at onset (from childhood to 81 years). While the pathogenic and likely pathogenic variants identified in this study can be used for genetic counseling, the VUS will require additional evidence, such as recurrence in unrelated patients, in order to be classified as pathogenic

Estimates of abundance and trend on a Blue Whale feeding ground off Isla de Chiloé, Chile

Since 1970, blue whales (Balaenoptera musculus) have been seen feeding in the waters off southern Chile during the summer and autumn (December to May). Investigation of the genetic, acoustic and morphological characteristics of these blue whales shows that they are a distinct but unnamed subspecies, called the Chilean blue whales. Photo-identification surveys have been conducted in the waters off northwestern Isla Grande de Chiloé, southern Chile from 2004–2012 and Isla Chañaral, central Chile in 2012. Over this time, 1,070 blue whales were encountered yielding, after photo-quality control, 318 and 267 unique photographs of the left and right side of the flank respectively. Using mark-recapture analysis of left and right side photographs collected from Isla Grande de Chiloé (2004–2012), open population models estimate that ~570–760 whales are feeding seasonally in this region. POPAN superpopulation abundance estimates for the same feeding ground in 2012 are 762 (95% confidence intervals, CI = 638–933) and 570 (95% CI 475–705) for left and right side datasets respectively, very similar to results from closed population models. Estimates of trend revealed strong variation in abundance, peaking in 2009 and [suggesting] fluctuating use in the survey area over time, likely related to the density of their prey. High inter-annual return rates suggest a degree of site-fidelity of individuals to Isla Grande de Chiloé and that the number of whales using this feeding ground is relatively small

Annotation of Genes Involved in Glycerolipid Biosynthesis in Chlamydomonas reinhardtii: Discovery of the Betaine Lipid Synthase BTA1(Cr)

Lipid metabolism in flowering plants has been intensely studied, and knowledge regarding the identities of genes encoding components of the major fatty acid and membrane lipid biosynthetic pathways is very extensive. We now present an in silico analysis of fatty acid and glycerolipid metabolism in an algal model, enabled by the recent availability of expressed sequence tag and genomic sequences of Chlamydomonas reinhardtii. Genes encoding proteins involved in membrane biogenesis were predicted on the basis of similarity to proteins with confirmed functions and were organized so as to reconstruct the major pathways of glycerolipid synthesis in Chlamydomonas. This analysis accounts for the majority of genes predicted to encode enzymes involved in anabolic reactions of membrane lipid biosynthesis and compares and contrasts these pathways in Chlamydomonas and flowering plants. As an important result of the bioinformatics analysis, we identified and isolated the C. reinhardtii BTA1 (BTA1(Cr)) gene and analyzed the bifunctional protein that it encodes; we predicted this protein to be sufficient for the synthesis of the betaine lipid diacylglyceryl-N,N,N-trimethylhomoserine (DGTS), a major membrane component in Chlamydomonas. Heterologous expression of BTA1(Cr) led to DGTS accumulation in Escherichia coli, which normally lacks this lipid, and allowed in vitro analysis of the enzymatic properties of BTA1(Cr). In contrast, in the bacterium Rhodobacter sphaeroides, two separate proteins, BtaA(Rs) and BtaB(Rs), are required for the biosynthesis of DGTS. Site-directed mutagenesis of the active sites of the two domains of BTA1(Cr) allowed us to study their activities separately, demonstrating directly their functional homology to the bacterial orthologs BtaA(Rs) and BtaB(Rs)