BioSciences

Colorectal cancer encompasses a heterogeneous group of malignancies that differ in pathophysiological mechanisms, immune response and infiltration, therapeutic response, and clinical prognosis. Numerous studies have highlighted the clinical relevance of tumor-infiltrating immune cells among different types of colorectal tumors yet vary in cell type definitions and cell identification strategies. The distinction of immune signatures is particularly challenging when several immune subtypes are involved but crucial to identify novel intercellular mechanisms within the tumor microenvironment. In this review, we compile human and non-human studies on tumor-infiltrating immune cells and provide an overview of immune subtypes, their pathophysiological functions, and their prognostic role in colorectal cancer. We discuss how differentiating immune signatures can guide the development of immunotherapeutic targets and personalized treatment regimens. We analyzed comprehensive human protein biomarker profiles across the entire immune spectrum to improve interpretability and application of tumor studies and to ultimately enhance immunotherapy and advance precision medicine for colorectal cancer patients.

Diterpenoid natural products serve critical functions in plant development and ecological adaptation and many diterpenoids have economic value as bioproducts. The family of class II diterpene synthases catalyzes the committed reactions in diterpenoid biosynthesis, converting a common geranylgeranyl diphosphate precursor into different bicyclic prenyl diphosphate scaffolds. Enzymatic rearrangement and modification of these precursors generate the diversity of bioactive diterpenoids. We report the crystal structure of Grindelia robusta 7,13-copalyl diphosphate synthase, GrTPS2, at 2.1 Å of resolution. GrTPS2 catalyzes the committed reaction in the biosynthesis of grindelic acid, which represents the signature metabolite in species of gumweed (Grindelia spp., Asteraceae). Grindelic acid has been explored as a potential source for drug leads and biofuel production. The GrTPS2 crystal structure adopts the conserved three-domain fold of class II diterpene synthases featuring a functional active site in the γβ-domain and a vestigial ɑ-domain. Substrate docking into the active site of the GrTPS2 apo protein structure predicted catalytic amino acids. Biochemical characterization of protein variants identified residues with impact on enzyme activity and catalytic specificity. Specifically, mutagenesis of Y457 provided mechanistic insight into the position-specific deprotonation of the intermediary carbocation to form the characteristic 7,13 double bond of 7,13-copalyl diphosphate.

Kenneth (Ken) Sauer was a mainstay of research in photosynthesis at the University of California, Berkeley and the Lawrence Berkeley National Laboratory (LBNL) for more than 50 years. Ken will be remembered by his colleagues, and other workers in the field of photosynthesis as well, for his pioneering work that introduced the physical techniques whose application have enriched our understanding of the basic reactions of oxygenic photosynthesis. His laboratory was a training ground for many students and postdocs who went on to success in the field of photosynthesis and many others. Trained as a physical chemist, he always brought that quantitative approach to research questions and used several spectroscopic methods in his research. His broad scientific interests concerned the role of manganese in oxygen evolution, electronic properties of chlorophylls, energy transport in antenna complexes, and electron transport reactions. He was also an enthusiastic teacher, an enormously successful mentor who leaves behind a legion of scientists as his abiding legacy, a lover of music and the outdoors with many interests beyond science, and a dedicated family man with a great sense of humility. In this tribute, we summarize some aspects of Ken Sauers life and career, illustrated with selected research achievements, and describe his approach to research and life as we perceived it, which is complemented by reminiscences of several current researchers in photosynthesis and other fields. The supporting material includes Ken Sauerss CV and publication list, as well as a list of the graduate students and postdocs he trained and of researchers that spent a sabbatical in his lab.

Photoprotection mechanisms are ubiquitous among photosynthetic organisms. The photoprotection capacity of the green alga Chlamydomonas reinhardtii is correlated with protein levels of stress-related light-harvesting complex (LHCSR) proteins, which are strongly induced by high light (HL). However, the dynamic response of overall thylakoid structure during acclimation to growth in HL has not been fully understood. Here, we combined live-cell super-resolution microscopy and analytical membrane subfractionation to investigate macroscale structural changes of thylakoid membranes during HL acclimation in Chlamydomonas. Subdiffraction-resolution live-cell imaging revealed that the overall thylakoid structures became thinned and shrunken during HL acclimation. The stromal space around the pyrenoid also became enlarged. Analytical density-dependent membrane fractionation indicated that the structural changes were partly a consequence of membrane unstacking. The analysis of both an LHCSR loss-of-function mutant, npq4 lhcsr1, and a regulatory mutant that over-expresses LHCSR, spa1-1, showed that structural changes occurred independently of LHCSR protein levels, demonstrating that LHCSR was neither necessary nor sufficient to induce the thylakoid structural changes associated with HL acclimation. In contrast, stt7-9, a mutant lacking a kinase of major light-harvesting antenna proteins, had a slower thylakoid structural response to HL relative to all other lines tested but still showed membrane unstacking. These results indicate that neither LHCSR- nor antenna-phosphorylation-dependent HL acclimation are required for the observed macroscale structural changes of thylakoid membranes in HL conditions.

During host infection, Toxoplasma gondii and related unicellular parasites move using gliding, which differs fundamentally from other known mechanisms of eukaryotic cell motility. Gliding is thought to be powered by a thin layer of flowing filamentous (F)-actin sandwiched between the plasma membrane and a myosin-covered inner membrane complex. How this surface actin layer drives the various gliding modes observed in experiments-helical, circular, twirling and patch, pendulum or rolling-is unclear. Here we suggest that F-actin flows arise through self-organization and develop a continuum model of emergent F-actin flow within the confines provided by Toxoplasma geometry. In the presence of F-actin turnover, our model predicts the emergence of a steady-state mode in which actin transport is largely directed rearward. Removing F-actin turnover leads to actin patches that recirculate up and down the cell, which we observe experimentally for drug-stabilized actin bundles in live Toxoplasma gondii parasites. These distinct self-organized actin states can account for observed gliding modes, illustrating how different forms of gliding motility can emerge as an intrinsic consequence of the self-organizing properties of F-actin flow in a confined geometry.

Organic bilayers or amorphous silica films of a few nanometer thickness featuring embedded molecular wires offer opportunities for chemically separating while at the same time electronically connecting photo- or electrocatalytic components. Such ultrathin membranes enable the integration of components for which direct coupling is not sufficiently efficient or stable. Photoelectrocatalytic systems for the generation or utilization of renewable energy are among the most prominent ones for which ultrathin separation layers open up new approaches for component integration for improving efficiency. Recent advances in the assembly and spectroscopic, microscopic, and photoelectrochemical characterization have enabled the systematic optimization of the structure, energetics, and density of embedded molecular wires for maximum charge transfer efficiency. The progress enables interfacial designs for the nanoscale integration of the incompatible oxidation and reduction catalysis environments of artificial photosystems and of microbial (or biomolecular)-abiotic systems for renewable energy.

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Zeaxanthin (Zea) is a key component in the energy-dependent, rapidly reversible, nonphotochemical quenching process (qE) that regulates photosynthetic light harvesting. Previous transient absorption (TA) studies suggested that Zea can participate in direct quenching via chlorophyll (Chl) to Zea energy transfer. However, the contamination of intrinsic exciton-exciton annihilation (EEA) makes the assignment of TA signal ambiguous. In this study, we present EEA-free TA data using Nicotiana benthamiana thylakoid membranes, including the wild type and three NPQ mutants (npq1, npq4, and lut2) generated by CRISPR/Cas9 mutagenesis. The results show a strong correlation between excitation energy transfer from excited Chl Qy to Zea S1 and the xanthophyll cycle during qE activation. Notably, a Lut S1 signal is absent in the npq1 thylakoids which lack zeaxanthin. Additionally, the fifth-order response analysis shows a reduction in the exciton diffusion length (LD) from 62 ± 6 nm to 43 ± 3 nm under high light illumination, consistent with the reduced range of exciton motion being a key aspect of plants response to excess light.

Photosynthesis-the conversion of energy from sunlight into chemical energy-is essential for life on Earth. Yet there is much we do not understand about photosynthetic energy conversion on a fundamental level: how it evolved and the extent of its diversity, its dynamics, and all the components and connections involved in its regulation. In this commentary, researchers working on fundamental aspects of photosynthesis including the light-dependent reactions, photorespiration, and C4 photosynthetic metabolism pose and discuss what they view as the most compelling open questions in their areas of research.