ABSTRACT
How proteins are targeted to lipid droplets (LDs) and distinguish the LD surface from the surfaces of other organelles is poorly understood, but many contain predicted amphipathic helices (AHs) that are involved in targeting. We have focused on human perilipin 4 (Plin4), which contains an AH that is exceptional in terms of length and repetitiveness. Using model cellular systems, we show that AH length, hydrophobicity, and charge are important for AH targeting to LDs and that these properties can compensate for one another, albeit at a loss of targeting specificity. Using synthetic lipids, we show that purified Plin4 AH binds poorly to lipid bilayers but strongly interacts with pure triglycerides, acting as a coat and forming small oil droplets. Because Plin4 overexpression alleviates LD instability under conditions where their coverage by phospholipids is limiting, we propose that the Plin4 AH replaces the LD lipid monolayer, for example during LD growth.
Subject(s)
Lipid Droplets/metabolism , Perilipin-4/chemistry , Perilipin-4/metabolism , Animals , Cell Line , Drosophila , HeLa Cells , Humans , Hydrophobic and Hydrophilic Interactions , Lipid Bilayers/chemistry , Lipid Bilayers/metabolism , Lipid Droplets/chemistry , Models, Molecular , Perilipin-4/genetics , Protein Binding , Protein Conformation, alpha-Helical , Protein Unfolding , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/metabolismABSTRACT
ResumenLas estimaciones de carbono en bosques tropicales, es muy importante para entender el papel de estos ecosistemas en el ciclo de este elemento. Este conocimiento permite apoyar y soportar las decisiones y la formulación de estrategias de mitigación y adaptación para reducir las emisiones de gases de efecto invernadero (GEI). Sin embargo, cuantificaciones detalladas de las existencias totales de carbono en los bosques montanos tropicales son limitadas, a pesar de su importancia científica y para el manejo de los ecosistemas (e.g. REDD+). Cuarenta y cuatro parcelas se establecieron en un paisaje heterogéneo compuesto por bosques maduros ubicados en el sur de los Andes colombianos. El objetivo fue identificar el papel de estos ecosistemas en el almacenamiento de carbono, evaluar la contribución de los reservorios analizados (biomasa aérea, biomasa subterránea y necromasa), y realizar contribuciones al enfoque de REDD+ en la escala de proyecto. En cada parcela se midieron todos los árboles, palmas y helechos con diámetro (D) ≥ 15 cm. En el caso de las palmas, se midió la altura de 40 % de los individuos, siguiendo el Protocolo Nacional de Colombia para estimar la biomasa y el carbono en los bosques naturales. Ecuaciones alométricas nacionales fueron utilizadas para estimar la biomasa aérea, y una ecuación general propuesta por el IPCC, fue utilizada para la estimación de la biomasa subterránea. La biomasa aérea de las palmas se estimó mediante un modelo local desarrollado para bosques montanos y premontanos. La necromasa se estimó para los árboles muertos en pie y los detritos gruesos. En este último caso, se midió la longitud y los diámetros de los extremos de las piezas. Las muestras para las estimaciones de densidad de madera, se recogieron en el campo y se analizaron en el laboratorio. El carbono promedio total se estimó en 545.9 ± 84.1 Mg/ ha (± SE). La biomasa aérea contribuyó con 72.5 %, la biomasa subterránea con 13.6 %, y la necromasa con 13.9 %. La principal conclusión de este estudio, es que los bosques montanos tropicales almacenan grandes cantidades de carbono, similares a las almacenadas por los bosques tropicales de tierras bajas. Además, se observó que la inclusión de otros compartimientos podría contribuir en más del 20 % de total del almacenamiento de carbono, lo que indica que las estimaciones que sólo incluyen la biomasa aérea, subestiman en gran medida las reservas de carbono en los ecosistemas forestales. Por tal razón, se sugiere incluir otros compartimientos en las estimaciones de carbono para iniciativas o proyectos REDD+.
AbstractCarbon estimations in tropical forests are very important to understand the role of these ecosystems in the carbon cycle, and to support decisions and the formulation of mitigation and adaptive strategies to reduce the greenhouse emission gases (GHG). Nevertheless, detailed ground-based quantifications of total carbon stocks in tropical montane forests are limited, despite their high value in science and ecosystem management (e.g. REDD+). The objective was to identify the role of these ecosystems as carbon stocks, to evaluate the contribution of the pools analyzed (aboveground biomass, belowground biomass and necromass), and to make contributions to the REDD+ approach from the project scale. For this study, we established 44 plots in a heterogeneous landscape composed by old-grown forests located in the Southern Colombian Andes. In each plot, all trees, palms and ferns with diameter (D) ≥ 15 cm were measured. In the case of palms, the height was measured for 40 % of the individuals, following the Colombia National Protocol to estimate biomass and carbon in natural forests. National allometric equations were used to estimate aboveground biomass, and a global equation proposed by IPCC was used for belowground biomass estimation; besides, palms' aboveground biomass was estimated using a local model. The necromass was estimated for dead standing trees and the gross debris. In the latter case, the length and diameters of the extremes in the pieces were measured. Samples for wood density estimations were collected in the field and analyzed in the laboratory. The mean total carbon stock was estimated as 545.9 ± 84.1 Mg/ha (± S.E.). The aboveground biomass contributed with 72.5 %, the belowground biomass with 13.6 %, and the necromass with 13.9 %. The main conclusion is that montane tropical forests store a huge amount of carbon, similar to low land tropical forests. In addition, the study found that the inclusion of other pools could contribute with more than 20 % to total carbon storage, indicating that estimates that only include the aboveground biomass, largely underestimate carbon stocks in tropical forest ecosystems. These results support the importance of including other carbon pools in REDD+ initiatives' estimations. Rev. Biol. Trop. 64 (1): 399-412. Epub 2016 March 01.
Subject(s)
Carbon/analysis , Forests , Biomass , Tropical Climate , ColombiaABSTRACT
The Saccharomyces cerevisiae Aft1 and Kluyveromyces lactis KlAft are orthologous yeast transcription activators that regulate the expression of the same group of iron-uptake genes but bind to the different DNA sites: TGCACCC for Aft1 and PuCACCC for KlAft. To establish whether the DNA-binding mechanisms of Aft1 and KlAft have diverged during the evolution of the Aft-type transcription factor, we examined the function of a nonconserved region in their DNA-binding domains. A large part of this region is composed of a sequence predicted to be disordered in structure and potentially phosphorylated. We show with deletion mutant analyses that this sequence is essential for the binding of Aft1 to its DNA site and for the iron uptake and growth of S. cerevisiae under iron-limited conditions. We constructed hybrid proteins by exchanging the nonconserved regions of Aft1 and KlAft. We show that the Aft1 region is necessary and sufficient for KlAft to bind efficiently to the Aft1 DNA site in S. cerevisiae and to complement the iron-dependent phenotype of the aft1Δaft2Δ mutant. This demonstrates that the changes in the nonconserved region of the Aft-type DNA-binding domain have led to changes in the DNA-binding specificity and have major consequences for the regulation of iron homeostasis. The combination of bioinformatic and experimental analyses indicates that the sequence TGCACCC is the most probable ancestral Aft-type element. Our findings suggest that the changes in the nonconserved region of the DNA-binding domain are responsible for the evolution of the TGCACCC sequence toward PuCACCC in the K. lactis species.
