Functional decreases in hydraulic and mechanical properties of field-grown transgenic poplar trees caused by modification of the lignin synthesis pathway through downregulation of the 4-coumarate:coenzyme A ligase gene

Society uses massive quantities of wood fiber in production of paper, and demand for fiber is projected to increase further as production of biofuels from fermentation of plant cellulosic materials increases. Because these end uses generally require the costly step of removing of lignin, wood with reduced or more easily extracted lignin has long been a goal of plant biotechnologists. However, there is little information on how reduction of lignin affects wood properties, tree development, and survival, especially in field environments. We studied a gene that had been previously reported to substantially reduce lignin content and improve biomass production in poplar. An antisense, xylem expressed version of the Pt4CL1 gene that encodes 4-coumarate:coenzyme A ligase (4CL) was inserted into hybrid poplar (Populus tremula x alba, INRA 717-1B4) and the growth and physiology of 14 transgenic lines (i.e., independent gene insertion events) was assessed over two growing seasons. Transgenic lines had 30-70% reductions in 4CL RNA expression in young shoots. This corresponded to 5-45% reduction in lignin as indicated by total monomer release through thioacidolysis and/or nitrobenzene oxidation. Only three transgenic lines with modest (>10%) reductions in lignin content sustained adequate growth and had normal tree form. Trees from five lines with severely reduced lignin formed up 24-60% of the stem cross-sectional area in brown colored wood that was essentially non-conductive to water, presumably due to the ectopic deposition of non-lignin phenolics and associated tyloses that occluded vessels. Across all genotypes, the transgenic lines had up to a three-fold increase in tensionwood, 40% lower modulus of elasticity, 25% lower modulus of rupture, 45% reduced resistance to xylem embolism (P₅₀), and a 60% increase in stem taper. Comparable patterns in wood density in lines that were lower in lignin by up to 9% by mass was compensated for by a 3% increase in polysaccharide content associated with tensionwood and a 6% increase in the deposition of extractives. Taken together, these data suggest that extensive field testing, ecophysiology, and wood quality evaluations are critical components of research and development on lignin-modified tree crops

Tree-ring Isotopes Adjacent to Lake Superior Reveal Cold Winter Anomalies for the Great Lakes Region of North America

Tree-ring carbon isotope discrimination (Δ13C) and oxygen isotopes (δ18O) collected from white pine (Pinus strobus) trees adjacent to Lake Superior show potential to produce the first winter-specific paleoclimate reconstruction with inter-annual resolution for this region. Isotopic signatures from 1976 to 2015 were strongly linked to antecedent winter minimum temperatures (Tmin), Lake Superior peak ice cover, and regional to continental-scale atmospheric winter pressure variability including the North American Dipole. The immense thermal inertia of Lake Superior underlies the unique connection between winter conditions and tree-ring Δ13C and δ18O signals from the following growing season in trees located near the lake. By combining these signals, we demonstrate feasibility to reconstruct variability in Tmin, ice cover, and continental-scale atmospheric circulation patterns (r ≥ 0.65, P \u3c 0.001)

Tyloses and phenolic deposits in xylem vessels impede water transport in low-lignin transgenic poplars: a study by cryo-fluorescence microscopy

Of 14 transgenic poplar genotypes (Populus tremula × Populus alba) with antisense 4-coumarate:coenzyme A ligase that were grown in the field for 2 years, five that had substantial lignin reductions also had greatly reduced xylem-specific conductivity compared with that of control trees and those transgenic events with small reductions in lignin. For the two events with the lowest xylem lignin contents (greater than 40% reduction), we used light microscopy methods and acid fuchsin dye ascent studies to clarify what caused their reduced transport efficiency. A novel protocol involving dye stabilization and cryo-fluorescence microscopy enabled us to visualize the dye at the cellular level and to identify water-conducting pathways in the xylem. Cryo-fixed branch segments were planed in the frozen state on a sliding cryo-microtome and observed with an epifluorescence microscope equipped with a cryo-stage. We could then distinguish clearly between phenolic-occluded vessels, conductive (stain-filled) vessels, and nonconductive (water- or gas-filled) vessels. Low-lignin trees contained areas of nonconductive, brown xylem with patches of collapsed cells and patches of noncollapsed cells filled with phenolics. In contrast, phenolics and nonconductive vessels were rarely observed in normal colored wood of the low-lignin events. The results of cryo-fluorescence light microscopy were supported by observations with a confocal microscope after freeze drying of cryo-planed samples. Moreover, after extraction of the phenolics, confocal microscopy revealed that many of the vessels in the nonconductive xylem were blocked with tyloses. We conclude that reduced transport efficiency of the transgenic low-lignin xylem was largely caused by blockages from tyloses and phenolic deposits within vessels rather than by xylem collapse

Spring temperature responses of oaks are synchronous with North Atlantic conditions during the last deglaciation

Paleoclimate proxies based on the measurement of xylem cell anatomy have rarely been developed across the temperature range of a species or applied to wood predating the most recent millennium. Here we describe wood anatomy-based proxies for spring temperatures in central North America from modern bur oaks (Quercus macrocarpa Michx.). The strong coherence of temperature signals across the species range supports the use of these proxies across thousands of years of climatic change. We also used 79 subfossil oak log cross sections from northern Missouri, ¹⁴C-dated to 9.9-13.63 ka (ka is 1000 cal yr BP), to assess the frequency of oak deposition into alluvial sediments and a subset of these oaks for a wood anatomy-based reconstruction of spring paleotemperatures. Temperatures during the Younger Dryas cold period (YD) were up to 3.5 degrees C lower than modern temperatures for that region, equivalent to or lower than those experienced at the northern edge of the modern species range. Compared to extant oaks growing at much higher [CO₂], subfossil oaks had greater vessel frequencies. Besides very low theoretical (or estimated) xylem conductivity near the beginning of the oak record near 13.6 ka, vessel frequencies greater than modern trees compensated for reduced vessel dimensions so that theoretical xylem conductivity was consistently above that of modern trees at the cold northern sites. Significant correlations were found between the frequency of ¹⁴C-dated oaks and either delta δ¹⁸O from the NGRIP (North Greenland Ice Core Project) ice core or from the Cariaco grayscale marine-sediment record from the southern Caribbean sea. Oak deposition into alluvial sediments during the YD was significantly lower than expected given the average sample depth of oaks from 9.9 to 13.6 ka. Reduced oak deposition during the YD suggests that an abrupt shift in climate reduced oak populations across the region and/or changed the rates of channel movement across drainages.Keywords: Pleistocene, Quercus macrocarpa, Holocene, Pre-Boreal, Younger Dryas, Radiocarbon, Wood anatomy, Great Plains\, USA, Phenology, Bolling-Allerod, Xylem, Bur oa

Seasonal and species‐level water‐use strategies and groundwater dependence in dryland riparian woodlands during extreme drought

Drought‐induced groundwater decline and warming associated with climate change are primary threats to dryland riparian woodlands. We used the extreme 2012–2019 drought in southern California as a natural experiment to assess how differences in water‐use strategies and groundwater dependence may influence the drought susceptibility of dryland riparian tree species with overlapping distributions. We analyzed tree‐ring stable carbon and oxygen isotopes collected from two cottonwood species (Populus trichocarpa and P. fremontii) along the semi‐arid Santa Clara River. We also modeled tree source water δ18O composition to compare with observed source water δ18O within the floodplain to infer patterns of groundwater reliance. Our results suggest that both species functioned as facultative phreatophytes that used shallow soil moisture when available but ultimately relied on groundwater to maintain physiological function during drought. We also observed apparent species differences in water‐use strategies and groundwater dependence related to their regional distributions. P. fremontii was constrained to more arid river segments and ostensibly used a greater proportion of groundwater to satisfy higher evaporative demand. P. fremontii maintained ∆13C at pre‐drought levels up until the peak of the drought, when trees experienced a precipitous decline in ∆13C. This response pattern suggests that trees prioritized maintaining photosynthetic processes over hydraulic safety, until a critical point. In contrast, P. trichocarpa showed a more gradual and sustained reduction in ∆13C, indicating that drought conditions induced stomatal closure and higher water use efficiency. This strategy may confer drought avoidance for P. trichocarpa while increasing its susceptibility to anticipated climate warming

Impacts of dwarf mistletoe on the physiology of host Tsuga heterophylla trees as recorded in tree-ring C and O stable isotopes

See article for Abstract. Keywords: mesophyll conductance, effective path lengt

Bacterial porin disrupts mitochondrial membrane potential and sensitizes host cells to apoptosis

The bacterial PorB porin, an ATP-binding beta-barrel protein of pathogenic Neisseria gonorrhoeae, triggers host cell apoptosis by an unknown mechanism. PorB is targeted to and imported by host cell mitochondria, causing the breakdown of the mitochondrial membrane potential (delta psi m). Here, we show that PorB induces the condensation of the mitochondrial matrix and the loss of cristae structures, sensitizing cells to the induction of apoptosis via signaling pathways activated by BH3-only proteins. PorB is imported into mitochondria through the general translocase TOM but, unexpectedly, is not recognized by the SAM sorting machinery, usually required for the assembly of beta-barrel proteins in the mitochondrial outer membrane. PorB integrates into the mitochondrial inner membrane, leading to the breakdown of delta psi m. The PorB channel is regulated by nucleotides and an isogenic PorB mutant defective in ATP-binding failed to induce delta psi m loss and apoptosis, demonstrating that dissipation of delta psi m is a requirement for cell death caused by neisserial infection

A dynamic leaf gas-exchange strategy is conserved in woody plants under changing ambient CO2: evidence from carbon isotope discrimination in paleo and CO2 enrichment studies

Rising atmospheric [CO2 ], ca , is expected to affect stomatal regulation of leaf gas-exchange of woody plants, thus influencing energy fluxes as well as carbon (C), water and nutrient cycling of forests. Researchers have proposed various strategies for stomatal regulation of leaf gas-exchange that include maintaining a constant leaf internal [CO2 ], ci , a constant drawdown in CO2 (ca - ci ), and a constant ci /ca . These strategies can result in drastically different consequences for leaf gas-exchange. The accuracy of Earth systems models depends in part on assumptions about generalizable patterns in leaf gas-exchange responses to varying ca . The concept of optimal stomatal behavior, exemplified by woody plants shifting along a continuum of these strategies, provides a unifying framework for understanding leaf gas-exchange responses to ca . To assess leaf gas-exchange regulation strategies, we analyzed patterns in ci inferred from studies reporting C stable isotope ratios (δ(13) C) or photosynthetic discrimination (∆) in woody angiosperms and gymnosperms that grew across a range of ca spanning at least 100 ppm. Our results suggest that much of the ca -induced changes in ci /ca occurred across ca spanning 200 to 400 ppm. These patterns imply that ca - ci will eventually approach a constant level at high ca because assimilation rates will reach a maximum and stomatal conductance of each species should be constrained to some minimum level. These analyses are not consistent with canalization towards any single strategy, particularly maintaining a constant ci . Rather, the results are consistent with the existence of a broadly conserved pattern of stomatal optimization in woody angiosperms and gymnosperms. This results in trees being profligate water users at low ca , when additional water loss is small for each unit of C gain, and increasingly water-conservative at high ca , when photosystems are saturated and water loss is large for each unit C gain. This article is protected by copyright. All rights reserved.Rising atmospheric [CO2], c(a), is expected to affect stomatal regulation of leaf gas-exchange of woody plants, thus influencing energy fluxes as well as carbon (C), water, and nutrient cycling of forests. Researchers have proposed various strategies for stomatal regulation of leaf gas-exchange that include maintaining a constant leaf internal [CO2], c(i), a constant drawdown in CO2 (c(a)-c(i)), and a constant c(i)/c(a). These strategies can result in drastically different consequences for leaf gas-exchange. The accuracy of Earth systems models depends in part on assumptions about generalizable patterns in leaf gas-exchange responses to varying c(a). The concept of optimal stomatal behavior, exemplified by woody plants shifting along a continuum of these strategies, provides a unifying framework for understanding leaf gas-exchange responses to c(a). To assess leaf gas-exchange regulation strategies, we analyzed patterns in c(i) inferred from studies reporting C stable isotope ratios (C-13) or photosynthetic discrimination () in woody angiosperms and gymnosperms that grew across a range of c(a) spanning at least 100ppm. Our results suggest that much of the c(a)-induced changes in c(i)/c(a) occurred across c(a) spanning 200 to 400ppm. These patterns imply that c(a)-c(i) will eventually approach a constant level at high c(a) because assimilation rates will reach a maximum and stomatal conductance of each species should be constrained to some minimum level. These analyses are not consistent with canalization toward any single strategy, particularly maintaining a constant c(i). Rather, the results are consistent with the existence of a broadly conserved pattern of stomatal optimization in woody angiosperms and gymnosperms. This results in trees being profligate water users at low c(a), when additional water loss is small for each unit of C gain, and increasingly water-conservative at high c(a), when photosystems are saturated and water loss is large for each unit C gain

Antisense down-regulation of 4CL expression alters lignification, tree growth and saccharification potential of field-grown poplar

Transgenic down-regulation of the Pt4CL1 gene family encoding 4-coumarate:coenzyme A ligase (4CL) has been reported as a means for reducing lignin content in cell walls and increasing overall growth rates, thereby improving feedstock quality for paper and bioethanol production. Using hybrid poplar (Populus tremula 3 Populus alba), we applied this strategy and examined field-grown transformants for both effects on wood biochemistry and tree productivity. The reductions in lignin contents obtained correlated well with 4CL RNA expression, with a sharp decrease in lignin amount being observed for RNA expression below approximately 50% of the nontransgenic control. Relatively small lignin reductions of approximately 10% were associated with reduced productivity, decreased wood syringyl/guaiacyl lignin monomer ratios, and a small increase in the level of incorporation of H-monomers (p-hydroxyphenyl) into cell walls. Transgenic events with less than approximately 50% 4CL RNA expression were characterized by patches of reddish-brown discolored wood that had approximately twice the extractive content of controls (largely complex polyphenolics). There was no evidence that substantially reduced lignin contents increased growth rates or saccharification potential. Our results suggest that the capacity for lignin reduction is limited; below a threshold, large changes in wood chemistry and plant metabolism were observed that adversely affected productivity and potential ethanol yield. They also underline the importance of field studies to obtain physiologically meaningful results and to support technology development with transgenic trees

Multi-ancestry GWAS of the electrocardiographic PR interval identifies 202 loci underlying cardiac conduction

The electrocardiographic PR interval reflects atrioventricular conduction, and is associated with conduction abnormalities, pacemaker implantation, atrial fibrillation (AF), and cardiovascular mortality. Here we report a multi-ancestry (N=293,051) genome-wide association meta-analysis for the PR interval, discovering 202 loci of which 141 have not previously been reported. Variants at identified loci increase the percentage of heritability explained, from 33.5% to 62.6%. We observe enrichment for cardiac muscle developmental/contractile and cytoskeletal genes, highlighting key regulation processes for atrioventricular conduction. Additionally, 8 loci not previously reported harbor genes underlying inherited arrhythmic syndromes and/or cardiomyopathies suggesting a role for these genes in cardiovascular pathology in the general population. We show that polygenic predisposition to PR interval duration is an endophenotype for cardiovascular disease, including distal conduction disease, AF, and atrioventricular pre-excitation. These findings advance our understanding of the polygenic basis of cardiac conduction, and the genetic relationship between PR interval duration and cardiovascular disease. On the electrocardiogram, the PR interval reflects conduction from the atria to ventricles and also serves as risk indicator of cardiovascular morbidity and mortality. Here, the authors perform genome-wide meta-analyses for PR interval in multiple ancestries and identify 141 previously unreported genetic loci.Peer reviewe