Larsen, J. B. | Search bioRxiv

bioRxiv2023

Comparative Phylogeography of Tree Species in the Hyrcanian Forests, Iran by Genome-wide SNPs

The Hyrcanian forests in Iran are one of the last remaining natural deciduous forests in the world and a UNESCO World Heritage site. We investigated the population genetics of representative indigenous tree species, Acer velutinum Boiss., Fagus orientalis Lipsky, and Quercus castaneifolia C.A. Mey. in northern Iran and also F. orientalis populations in the Euro-Siberian and Colchic sub-regions in northern Turkiye. We used the double-digest RAD-seq method and genotyped 90 populations and 1,589 individuals across the distribution range of the species. Our analyses yielded 1,347, 2,091, and 8,881 genome-wide SNPs from 28 populations of A. velutinum, 32 populations of F. orientalis, and 30 Q. castaneifolia, respectively. Our results revealed higher genetic differentiations among A. velutinum populations than those of F. orientalis and Q. castaneifolia within the Hyrcanian forests. The global FST value was lowest for F. orientalis populations (0.019) and highest for A. velutinum populations (0.12), while the FIS value was negative for A. velutinum (-0.095). Demographic history analysis indicated a bottleneck during the last glacial maximum for the A. velutinum populations with reduced effective population size. The three species showed evidence of population bottlenecks during the Pliocene. Our findings highlight the pronounced genetic divergence among A. velutinum populations in the Hyrcanian forests compared to the other two species, suggesting cryptic speciation. Conversely, F. orientalis and Q. castaneifolia populations demonstrated a reduced level of genetic structure, indicating that species-specific factors, such as pollen production and pollination efficiency, may have influenced the genetic patterns within these species in similar environments. We did not observe significant elevational genetic differentiation within populations of the studied species. Furthermore, the F. orientalis populations from Turkiye exhibited a distinct west-east genetic structure and were highly diverged from the Iranian F. orientalis populations.

evolutionary biology↗

bioRxiv2023

Tuning the double lipidation of salmon calcitonin to introduce a pore-like membrane translocation mechanism

A widespread strategy to increase the transport of therapeutic peptides across cellular membranes has been to attach lipid moieties to the peptide backbone (lipidation) to enhance their intrinsic membrane interaction. Efforts in vitro and in vivo investigating the correlation between lipidation characteristics and peptide membrane translocation efficiency have traditionally relied on end-point read-out assays and trial-and-error-based optimization strategies. Consequently, the molecular details of how therapeutic peptide lipidation affects its membrane permeation and translocation mechanisms remain unresolved. Here we employed salmon calcitonin as a model therapeutic peptide and synthesized nine double lipidated analogs with varying lipid chain lengths. We used single giant unilamellar vesicle (GUV) calcein influx time-lapse fluorescence microscopy to determine how tuning the lipidation length can lead to an All-or-None GUV filling mechanism, indicative of a peptide mediated pore formation. Finally, we used a GUVs-containing-inner-GUVs assay to demonstrate that only peptide analogs capable of inducing pore formation show efficient membrane translocation. Our data provided the first mechanistic details on how therapeutic peptide lipidation affects their membrane perturbation mechanism and demonstrated that fine-tuning lipidation parameters could induce an intrinsic pore-forming capability. These insights and the microscopy based workflow introduced for investigating structure-function relations could be pivotal for optimizing future peptide design strategies. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=197 SRC="FIGDIR/small/548826v1_ufig1.gif" ALT="Figure 1"> View larger version (87K): org.highwire.dtl.DTLVardef@23430org.highwire.dtl.DTLVardef@b06145org.highwire.dtl.DTLVardef@1c22579org.highwire.dtl.DTLVardef@12a56eb_HPS_FORMAT_FIGEXP M_FIG C_FIG Highlights- Lipidating the therapeutic peptide salmon calcitonin alters its biophysical characteristics, including oligomer size, hydrophobicity and membrane activity. - Fluorescent microscopy of single GUVs enables the determination of peptide mediated reporter dye influx behavior as either graded or All-or-None, which is coupled to either smaller membrane perturbations or peptide pore formation. - Modulating the number of hydrocarbons constituting the lipidation moieties determines the membrane permeation mechanism. - By increasing the lipid chain length lipidated of salmon calcitonin goes from displaying smaller membrane perturbations to a peptide pore formation mechanism. - Effective membrane translocation of lipidated salmon calcitonin requires a peptide mediated pore forming mechanism.

biophysics↗