bluuebunny/crossref_metadata_2025_split huggingface dataset:
The cytoplasmic tail of myelin protein zero induces morphological changes in lipid membranes
Authors: Krokengen Oda C. , Touma Christine , Mularski Anna , Sutinen Aleksi , Dunkel Ryan , Ytterdal Marie , Raasakka Arne , Mertens Haydyn D.T. , Simonsen Adam Cohen , Kursula Petri 
Published: March 2024

AbstractThe major myelin protein expressed by the peripheral nervous system Schwann cells is protein zero (P0), representing 50% of the total protein content in myelin. This 30-kDa integral membrane protein consists of an immunoglobulin (Ig)-like domain, a transmembrane helix, and a 69-residue C-terminal cytoplasmic tail (P0ct). The basic residues in P0ct contribute to the tight packing of the myelin lipid bilayers, and alterations in the tail affect how P0 functions as an adhesion molecule necessary for the stability of compact myelin. Several neurodegenerative neuropathies are related to P0, including the more common Charcot-Marie-Tooth disease (CMT) and Dejerine-Sottas syndrome (DSS), but also rare cases of motor and sensory polyneuropathy. We find that high P0ct concentrations affect the membrane properties of bicelles and induce a lamellar-to-inverted hexagonal phase transition, which causes the bicelles to fuse into long, protein-containing filament-like structures. These structures likely reflect the formation of semi-crystalline lipid domains of potential relevance for myelination. Not only is P0ct important for stacking lipid membranes, but time-lapse fluorescence microscopy shows that it might affect membrane properties during myelination. We further describe recombinant production and low-resolution structural characterization of full-length human P0. Our findings shed light on P0ct effects on membrane properties, and with successful purification of full-length P0, we have new tools to studyin vitrothe role P0 has in myelin formation and maintenance.
MOSAIC: a highly efficient, one-step recombineering approach to plasmid editing and diversification
Authors: van den Brink Marijn , Althuis Timotheus Y. , Danelon Christophe , Claassens Nico J. 
Published: March 2024

ABSTRACTThe editing of plasmids and construction of plasmid libraries is paramount to the engineering of desired functionalities in synthetic biology. Typically, plasmids with targeted mutations are produced through time– and resource-consuming DNA amplification and/or cloning steps. In this study, we establish MOSAIC, a highly efficient protocol for the editing of plasmids and generation of combinatorial plasmid libraries. This quick protocol employs the efficient single-stranded DNA annealing protein (SSAP) CspRecT to incorporate (libraries of) DNA oligos harboring the desired mutations into a target plasmid inE. coli.In addition to up to 90% single-target plasmid editing efficiency, we demonstrate that MOSAIC enables the generation of a combinatorial plasmid library spanning four different target regions on a plasmid, in a single transformation. Lastly, we integrated a user-friendly validation pipeline using Nanopore sequencing reads, requiring minimal computational experience. We anticipate that MOSAIC will provide researchers with a simple, rapid and resource-effective method to edit plasmids or generate large, diverse plasmid libraries for a wide range ofin vivoorin vitroapplications in molecular and synthetic biology.Abstract FigureGRAPHICAL ABSTRACT
Aeromonas hydrophilaCobQ is a new type of NAD+- and Zn2+-independent protein lysine deacetylase
Authors: Wang Yuqian , Wang Guibin , Zhang Lishan , Cai Qilan , Lin Meizhen , Huang Dongping , Xie Yuyue , Lin Wenxiong , Lin Xiangmin 
Published: March 2024

AbstractProtein N−-lysine acetylation (Kac) modifications play crucial roles in diverse physiological and pathological functions in cells. In prokaryotic cells, there are only two types of lysine deacetylases (KDACs) that are Zn2+- or NAD+-dependent. In this study, we reported a protein, AhCobQ, inAeromonas hydrophilaATCC 7966 that presents NAD+- and Zn2+-independent KDAC activity. Furthermore, its KDAC activity is located in an unidentified domain (from 195–245 aa). Interestingly, AhCobQ has no homology with current known KDACs, and no homologous protein was found in eukaryotic cells. A protein substrate analysis showed that AhCobQ has specific protein substrates in common with other known KDACs, indicating that these KDACs can dynamically co-regulate the states of Kac proteins. Microbiological methods employed in this study affirmed AhCobQ’s positive regulation of isocitrate dehydrogenase (ICD) enzymatic activity at the K388 site, implicating AhCobQ in the modulation of bacterial enzymatic activities. In summary, our findings present compelling evidence that AhCobQ represents a distinctive type of KDAC with significant roles in bacterial biological functions.SummaryThe lack of exploration of new lysine acetylases and deacetylases (KDACs) and their protein substrates in prokaryotic cells has become a bottleneck in the functional study of lysine acetylation modifications. In this study, we reported a novel Zn2+- and NAD+-independent KDAC protein, AhCobQ, inAeromonas hydrophila. Interestingly, this protein does not share homology with current known KDACs, and its KDAC activity is located in an unknown domain for which a homologous protein cannot be found in eukaryotic cells. The following analysis showed that AhCobQ affected the enzymatic activity and protein-protein interaction ability of its protein substrates. In summary, these results extended our understanding of the regulatory mechanism of bacterial lysine acetylation modifications.HighlightsAhCobQ is an NAD+- and Zn2+-independent protein lysine deacetylase.There are no proteins homologous to AhCobQ in eukaryotes.The deacetylase activity of AhCobQ is located in an unknown domain.AhCobQ has specific protein substrates and substrates in common with other lysine deacetylases.AhCobQ positively regulates the enzymatic activity of isocitrate dehydrogenase at its K388 site.
Interactions betweenPseudomonas aeruginosaand six opportunistic pathogens cover a broad spectrum from mutualism to antagonism
Authors: Laffont Clémentine , Wechsler Tobias , Kümmerli Rolf 
Published: March 2024

AbstractBacterial infections often involve more than one pathogen. While it is known that polymicrobial infections can impact disease outcomes, we have a poor understanding about how pathogens affect each other’s behaviour and fitness. Here, we used a microscopy approach to explore interactions betweenPseudomonas aeruginosaand six opportunistic human pathogens that often co-occur in polymicrobial infections:Acinetobacter baumannii,Burkholderia cenocepacia,Escherichia coli,Enterococcus faecium,Klebsiella pneumoniae, andStaphylococcus aureus.When following growing micro-colonies on agarose pads over time, we observed a broad spectrum of species-specific ecological interactions, ranging from mutualism to antagonism. For example,P. aeruginosaengaged in a mutually beneficial interaction withE. faeciumbut suffered from antagonism byE. coliandK. pneumoniae. While we found little evidence for active directional growth towards or away from cohabitants, we observed that certain species increased growth in double layers in co-cultures and that physical forces due to fast colony expansion had a major impact on fitness and interaction patterns. Overall, our work provides an atlas of pathogen interactions, potentially useful to understand species dynamics in polymicrobial infections. We discuss possible mechanisms driving pathogen interactions and offer predictions of how the different ecological interactions could affect virulence.
Adaptive evolution of sesquiterpene deoxyphomenone in mycoparasitism byHansfordia pulvinataassociated with horizontal gene transfer fromAspergillusspecies
Authors: Maeda Kazuya , Sumita Takuya , Nishi Oumi , Sushida Hirotoshi , Higashi Yumiko , Nakagawa Hiroyuki , Suzuki Tomoko , Iwao Eishin , Fanani Much Zaenal , Nishiya Yoshiaki , Iida Yuichiro 
Published: March 2024

AbstractLeaf mold caused by the ascomycete fungusCladosporium fulvumis a devastating disease of tomato plants. The mycoparasitic fungusHansfordia pulvinatais an effective biocontrol agent that parasitizesC. fulvumhyphae on leaves and secretes 13-deoxyphomenone, an eremophilane-type sesquiterpene, which was also identified as a sporulation-inducing factor inAspergillus oryzae. Here, we identified deoxyphomenone biosynthesis (DPH) gene clusters conserved in bothH. pulvinataandAspergillussectionFlaviincludingA. oryzaeandA. flavus. Functional disruption ofDPH1orthologous genes encoding sesquiterpene cyclase inH. pulvinata,A. oryzaeand its close relativeA. flavusrevealed that deoxyphomenone inH. pulvinatahad exogenic antifungal activity against the host fungusC. fulvumand controlled endogenic sporulation inAspergillusspecies. Deoxyphomenone also inhibited mycelial growth ofC. fulvumand the non-host tomato pathogenPseudocercospora fuligena. CompleteDPHclusters, highly similar to those inH. pulvinata, were exclusive toAspergillussectionFlavi, while species in otherAspergillussections contained fragmentedDPHclusters. A comparative genomics analysis revealed that theseDPHgene clusters share a common origin and are horizontally transferred across large taxonomic distances from an ancestor ofAspergillustoH. pulvinata. Our results suggest that, after horizontal transfer,H. pulvinatamaintained theDPHcluster as the inhibitory effect of deoxyphomenone on spore germination and mycelial growth contributed to its mycoparasitism on the host fungusC. fulvum.
Mar1, an HMG-box protein, regulatesn-alkane adsorption and cell morphology of the dimorphic yeastYarrowia lipolytica
Authors: Kimura-Ishimaru Chiaki , Liang Simiao , Matsuse Katsuro , Iwama Ryo , Sato Kenta , Watanabe Natsuhito , Tezaki Satoshi , Horiuchi Hiroyuki , Fukuda Ryouichi 
Published: March 2024

ABSTRACTThe dimorphic yeastYarrowia lipolyticapossesses an excellent ability to utilizen-alkane as a sole carbon and energy source. Although there are detailed studies on the enzymes that catalyze the reactions in the metabolic processes ofn-alkane inY. lipolytica, the molecular mechanism underlying the incorporation ofn-alkane into the cells remains to be elucidated. BecauseY. lipolyticaadsorbsn-alkane, we postulated thatY. lipolyticaincorporatesn-alkane through direct interaction with it. We isolated and characterized mutants defective in adsorption ton-hexadecane. One of the mutants harbored a nonsense mutation inMAR1(Morphology andn-alkaneAdsorptionRegulator) encoding a protein containing a high mobility group box. The deletion mutant ofMAR1exhibited defects in adsorption ton-hexadecane and filamentous growth on solid media, whereas the strain that overexpressedMAR1exhibited hyperfilamentous growth. Fluorescence microscopic observations suggested that Mar1 localizes in the nucleus. RNA-seq analysis revealed the alteration of the transcript levels of several genes, including those encoding transcription factors and cell surface proteins, by the deletion ofMAR1. These findings suggest thatMAR1is involved in the transcriptional regulation of the genes required forn-alkane adsorption and cell morphology transition.IMPORTANCEY. lipolytica, a dimorphic yeast capable of assimilatingn-alkane as a carbon and energy source, has been extensively studied as a promising host for bioconversion ofn-alkane into useful chemicals and bioremediation of soil and water contaminated by petroleum. While the metabolic pathway ofn-alkane in this yeast and the enzymes involved in this pathway have been well-characterized, the molecular mechanism to incorporaten-alkane into the cells is yet to be fully understood. Due to the ability ofY. lipolyticato adsorbs ton-alkane, it has been hypothesized thatY. lipolyticaincorporatesn-alkane through direct interaction with it. In this study, we identified a gene,MAR1, which plays a crucial role in the transcriptional regulation of the genes necessary for the adsorption ton-alkane and the transition of the cell morphology inY. lipolytica. Our findings provide valuable insights that could lead to advanced applications ofY. lipolyticainn-alkane bioconversion and bioremediation.
Characterization of novel role for Rab27B in autophagy regulation in colorectal cancer
Authors: Afroz Sahida , Preet Ranjan , Vishwakarma Vikalp , Evans Andrew E. , Dixon Dan A. 
Published: March 2024

ABSTRACTIntroductionAutophagy is a dynamic, multi-step process that cells use to degrade damaged, abnormal, and potentially harmful cellular substances. While autophagy is maintained at a basal level in all cells, it is activated at a higher level in many cancer cells and promotes tumor growth, anti-tumor immune response, and resistance to cancer therapy. As a result, autophagy is increasingly being recognized to have an important role in cancer progression and emerging as a potential target for cancer therapy. We recently discovered that small GTPase Rab27B, a known regulator of vesicle trafficking and exosome secretion, is also involved in the autophagy process.MethodsRab27B was knocked out using CRISPR/Cas9 in CRC cell line HCT116. Western blotting, Immunofluorescence, MTT assay, spheroid formation assay, soft agar assay and xenograft studies were performed to analyze the effects of Rab27B deletion on CRC cells.ResultsCRISPR/Cas9 deletion or siRNA knockdown of Rab27B in colorectal cancer cells (CRC) showed an abnormal accumulation of autophagy vesicles. Additionally, we observed a significant increase in the autophagy markers LC3-II and p62 by immunocytochemistry and western blot analysis, suggesting a defect in the autophagy flux process. Lysotracker and mCherry-EGFP-LC3 fusion construct indicate an impairment in autophagosome and lysosome fusion when Rab27B is silenced. This defect was rescued by full-length and constitutively active GTP mutant of Rab27B. As autophagy has been shown to have a pro-survival role in tumor growth and stress response, we hypothesized that the observed defects in autophagy flux resulting from Rab27B loss would cause reduced stress response and tumor growth. Indeed, Rab27B knockout reduced cell viability in response to starvation and a 94% reduction in soft agar colony formation. Rab27B deletion also prevented spheroid formationin vitro.Finally, to analyze the effect of Rab27B deletion in tumor formationin vivo, we performed a xenograft study with wildtype and Rab27B knockout CRC cells, resulting in a dramatic loss of tumor growth (p<0.0001) in the KO cells.ConclusionsTogether, our results demonstrate a new role of Rab27B in the autophagy trafficking process in CRC. Future studies will focus on investigating the mechanism of how Rab27B functions in the autophagy pathway and whether Rab27B can be targeted as a potential therapeutic strategy for CRC.
Discovery of a cell-active chikungunya virus nsP2 protease inhibitor using a covalent fragment-based screening approach
Authors: Merten Eric M. , Sears John D. , Leisner Tina M. , Hardy Paul B. , Ghoshal Anirban , Hossain Mohammad Anwar , Asressu Kesatebrhan Haile , Brown Peter J. , Stashko Michael A. , Herring Laura , Mordant Angie L. , Webb Thomas S. , Mills Christine A. , Barker Natalie K. , Streblow Zachary J. , Perveen Sumera , Arrowsmith Cheryl , Arnold Jamie J. , Cameron Craig E , Streblow Daniel N. , Moorman Nathaniel J. , Heise Mark , Willson Timothy M. , Popov Konstantin , Pearce Kenneth H. 
Published: March 2024

AbstractChikungunya virus (CHIKV) is a mosquito-borne alphavirus that has been responsible for numerous large-scale outbreaks in the last twenty years. Currently, there are no FDA-approved therapeutics for any alphavirus infection. CHIKV non-structural protein 2 (nsP2), which contains a cysteine protease domain, is essential for viral replication, making it an attractive target for a drug discovery campaign. Here, we optimized a CHIKV nsP2 protease (nsP2pro) biochemical assay for the screening of a 6,120-compound cysteine-directed covalent fragment library. Using a 50% inhibition threshold, we identified 153 hits (2.5% hit rate). In dose-response follow up, RA-0002034, a covalent fragment that contains a vinyl sulfone warhead, inhibited CHIKV nsP2pro with an IC50of 58 ± 17 nM, and further analysis with time-dependent inhibition studies yielded a kinact/KIof 6.4 x 103M-1s-1. LC-MS/MS analysis determined that RA-0002034 covalently modified the catalytic cysteine in a site-specific manner. Additionally, RA-0002034 showed no significant off-target reactivity against a panel of cysteine proteases. In addition to the potent biochemical inhibition of CHIKV nsP2pro activity and exceptional selectivity, RA-0002034 was tested in cellular models of alphavirus infection and effectively inhibited viral replication of both CHIKV and related alphaviruses. This study highlights the discovery and characterization of the chemical probe RA-0002034 as a promising hit compound from covalent fragment-based screening for development toward a CHIKV or pan-alphavirus therapeutic.Significance StatementChikungunya virus is one of the most prominent and widespread alphaviruses and has caused explosive outbreaks of arthritic disease. Currently, there are no FDA-approved drugs to treat disease caused by chikungunya virus or any other alphavirus-caused infection. Here, we report the discovery of a covalent small molecule inhibitor of chikungunya virus nsP2 protease activity and viral replication of four diverse alphaviruses. This finding highlights the utility of covalent fragment screening for inhibitor discovery and represents a starting point towards the development of alphavirus therapeutics targeting nsP2 protease.
Does relationship conflict reduce novel idea communication through perceived leader openness? Power distance orientation as a moderator
Authors: Tsai Ming-Hong 
Published: March 2024

Purpose
This paper aims to investigate why followers have low perceptions of leader openness and thus feel reluctant to communicate novel ideas by examining leader–follower relationship conflict (i.e. interpersonal incompatibility) and a follower’s power distance orientation (i.e. an acceptance of uneven power distribution in organizations) as antecedents.

Design/methodology/approach
The research administrators conducted a three-wave work behavior survey in Study 1, a laboratory experiment in Study 2, and an online experiment in Study 3.

Findings
The results demonstrated that leader–follower relationship conflict reduced followers’ perceptions of leader openness. However, the negative impact of relationship conflict became non-significant when followers have high power distance orientations (i.e. an acceptance of uneven power distribution in organizations). The findings also showed an indirect interaction effect of leader–follower relationship conflict and followers’ power distance orientation on the followers’ communication of novel ideas through the followers’ perceptions of leader openness.

Originality/value
The research suggests that followers with higher power distance orientations are more likely to communicate novel ideas consistently because their relationship conflicts with their leaders do not negatively influence their perceptions of leader openness. Although researchers traditionally view cultures with a high level of power distance value as an obstacle to employee creativity, the present study reveals the benefits of an individual-level power distance orientation.
Identification of key bottlenecks in human rights-based approach to family planning among HIV-infected women in a resource-restricted setting of Nepal
Authors: Dhungana Govinda Prasad , Bhatta Dwij Raj , Zhang Wei-Hong 
Published: March 2024

Purpose
Family planning (FP) services through the lens of human rights are not well known in Nepal. This study aims to assess Family Planning 2020: Rights and Empowerment Principles for Family Planning and identify factors affecting contraceptive use among HIV-infected women living in rural Far Western Nepal.

Design/methodology/approach
This study conducted a cross-sectional survey using self-designed proforma. To assess the association between contraceptive use and independent variables, this study calculated adjusted odd ratio (AOR) with 95% confidence interval (CI) using statistical package for social sciences (SPSS) V.20.

Findings
Only 37.8% of participants had access to full range of contraceptive methods, and only 57.5% of participants received proper counseling. Agency/autonomy, transparency/accountability and voice/participation were practiced by 43.7%, 23.4% and 19.7% of participants, respectively. Husband’s support (AOR = 4.263; 95% CI: 1.640–11.086), availability of FP services in their locality (AOR = 2.497; 95% CI: 1.311–4.754), employment (AOR = 3.499; 95% CI: 1.186–10.328) and postpartum period (AOR = 0.103; 95% CI: 0.023–0.475) were significantly associated with contraceptive use.

Research limitations/implications
Health-care providers’ and program managers’ perspectives were not examined.

Practical implications
Findings of this study will be useful for making strategic plan on human rights-based approach to FP.

Social implications
Expanding access to contraceptive information and services and strengthening autonomy, accountability and participation are key to human rights-based approach to FP.

Originality/value
This study identified that inadequate counselling, nonavailability of full range of contraceptive methods, low level of autonomy, accountability and participation were key bottlenecks in fulfilling human rights-based approach to FP.