Identification of cellular signatures associated with chinese hamster ovary cell adaptation for secretion of antibodies.

The secretory capacity of Chinese hamster ovary (CHO) cells remains a fundamental bottleneck in the manufacturing of protein-based therapeutics. Unconventional biological drugs with complex structures and processing requirements are particularly problematic. Although engineered vector DNA elements can achieve rapid and high-level therapeutic protein production, a high metabolic and protein folding burden is imposed on the host cell.

Expansion of the Structure-Activity Relationship Profile of Triaminopyrimidines as Inhibitors of Caspase-1.

Caspase-1 is a sought-after therapeutic target for inflammatory conditions due to its role in activation and release of pro-inflammatory cytokines, but there has been little success getting drugs into the clinic. We have previously shown triaminopyrimidines such as CK-1-41 are potent, reversible small molecule inhibitors of caspase-1, likely binding in an allosteric site within the enzyme. A series of analogs of CK-1-41 were synthesized and tested against caspase-1 to develop a more robust structure-activity relationship profile.

Balanced parenting: Proposing a differentiation-based parenting approach informed by Bowen family systems theory.

The purpose of this article is to introduce Balanced Parenting, a differentiation-based parenting approach informed by Bowen family systems theory. It is aimed at countering today's anxious, child-focused parenting culture and deconstructing attachment theory, which has been identified as contributing to this culture. The approach includes four main pillars: (a) deconstructing attachment theory, (b) the five principles of balanced parenting, (c) shifting the paradigm, and (d) steps toward change.

The Minnesota Critical Care Working Group 1: Monitoring and coordinating statewide critical care surge response in the COVID-19 pandemic, March 2020 through July 1, 2021.

Background: In response to the COVID-19 pandemic and as part of the statewide healthcare coalition response the Minnesota Critical Care Working Group (CCWG), composed of Interprofessional leaders from the state's nine largest health systems was established and entrusted to plan and coordinate critical care support for Minnesota from March 2020 through July1, 2021.

Research Question: Can a statewide Critical Care Working Group develop contingency and crisis level surge strategies and indicators in response to the COVID-19 pandemic while evolving into a highly collaborative team?

Study Design And Methods: CCWG members (Intensivists, ethicists, nurses, MDH and MHA leaders) met by audio video conferencing as often as daily assessing COVID and non-COVID hospitalization data, developed surge indicators reflecting contingency versus crisis conditions, and planned responses collaboratively. A foundation of collaboration and teamwork developed which facilitated an effective statewide response.

The Minnesota Critical Care Working Group 2: Crisis conditions during the COVID-19 Pandemic, July 2021 through March 2022.

Background: At the request of the Statewide Healthcare Coordination Center, the Minnesota Critical Care Working Group (CCWG) and ethics subgroup (EWG), composed of interprofessional leaders from Minnesota's nine largest health systems were asked to plan and coordinate critical care operations during the COVID-19 pandemic, including the 2021 Fall surge.

Research Question: Can a statewide Working Group collaboratively analyze real time evidence to identify crisis conditions and engage state leadership to implement care processes?

Study Design And Methods: CCWG/EWG met via video conferencing during the Fall 2021 severe surge to analyze evidence and plan for potential crisis care conditions. Five sources of evidence informed their actions including group consensus on operating conditions; Federal Tele-Tracking data; MOCC patient placement data; and two surveys created and distributed to hospitals and healthcare professionals.

ER export via SURF4 uses diverse mechanisms of both client and coat engagement.

Protein secretion is an essential process that drives cell growth and communication. Enrichment of soluble secretory proteins into ER-derived transport carriers occurs via transmembrane cargo receptors that connect lumenal cargo to the cytosolic COPII coat. Here, we find that the cargo receptor, SURF4, recruits different SEC24 cargo adaptor paralogs of the COPII coat to export different cargoes.

Cryo-electron tomography reveals how COPII assembles on cargo-containing membranes.

Proteins traverse the eukaryotic secretory pathway through membrane trafficking between organelles. The coat protein complex II (COPII) mediates the anterograde transport of newly synthesized proteins from the endoplasmic reticulum, engaging cargoes with a wide range of size and biophysical properties. The native architecture of the COPII coat and how cargo might influence COPII carrier morphology remain poorly understood.

Survey of antimicrobial and probiotic use practices in wildlife rehabilitation in the United States.

Antimicrobial resistance is a global health concern. As such, there have been increased efforts to monitor and standardize antimicrobial prescribing practices in humans and domestic animals. In contrast, there is relatively little known about specific prescribing practices in wild animals despite the wide use of antimicrobials and other microbial interventions, such as probiotics to treat captive wildlife.

SNARE chaperone Sly1 directly mediates close-range vesicle tethering.

The essential Golgi protein Sly1 is a member of the Sec1/mammalian Unc-18 (SM) family of SNARE chaperones. Sly1 was originally identified through remarkable gain-of-function alleles that bypass requirements for diverse vesicle tethering factors. Employing genetic analyses and chemically defined reconstitutions of ER-Golgi fusion, we discovered that a loop conserved among Sly1 family members is not only autoinhibitory but also acts as a positive effector.

Dynamic conformational changes of a tardigrade group-3 late embryogenesis abundant protein modulate membrane biophysical properties.

A number of intrinsically disordered proteins (IDPs) encoded in stress-tolerant organisms, such as tardigrade, can confer fitness advantage and abiotic stress tolerance when heterologously expressed. Tardigrade-specific disordered proteins including the cytosolic-abundant heat-soluble proteins are proposed to confer stress tolerance through vitrification or gelation, whereas evolutionarily conserved IDPs in tardigrades may contribute to stress tolerance through other biophysical mechanisms. In this study, we characterized the mechanism of action of an evolutionarily conserved, tardigrade IDP, HeLEA1, which belongs to the group-3 late embryogenesis abundant (LEA) protein family.

In situ architecture of the ER-mitochondria encounter structure.

The endoplasmic reticulum and mitochondria are main hubs of eukaryotic membrane biogenesis that rely on lipid exchange via membrane contact sites, but the underpinning mechanisms remain poorly understood. In yeast, tethering and lipid transfer between the two organelles is mediated by the endoplasmic reticulum-mitochondria encounter structure (ERMES), a four-subunit complex of unresolved stoichiometry and architecture. Here we determined the molecular organization of ERMES within Saccharomyces cerevisiae cells using integrative structural biology by combining quantitative live imaging, cryo-correlative microscopy, subtomogram averaging and molecular modelling.

Survey of clinical and commensal Escherichia coli from commercial broilers and turkeys, with emphasis on high-risk clones using APECTyper.

Molecular characterization of avian pathogenic Escherichia coli (APEC) is challenging due to the complex nature of its associated disease, colibacillosis, in poultry. Numerous efforts have been made toward defining APEC, and it is becoming clear that certain clonal backgrounds are predictive of an avian E. coli isolate's virulence potential.

Effects of a postbiotic, with and without a saponin-based product, on turkey performance.

Modern poultry production relies on an ability to prevent and mitigate challenges to bird health, while maintaining a productive bird. A number of different classes of biologics-based feed additives exist, and many have been tested individually for their impacts on poultry health and performance. Fewer studies have examined the combinations of different classes of products.

Yeast Svf1 binds ceramides and contributes to sphingolipid metabolism at the ER cis-Golgi interface.

Ceramides are essential precursors of complex sphingolipids and act as potent signaling molecules. Ceramides are synthesized in the endoplasmic reticulum (ER) and receive their head-groups in the Golgi apparatus, yielding complex sphingolipids (SPs). Transport of ceramides between the ER and the Golgi is executed by the essential ceramide transport protein (CERT) in mammalian cells.

Ultrastructure of COPII vesicle formation in yeast characterized by correlative light and electron microscopy.

Traffic of proteins out of the endoplasmic reticulum (ER) is driven by the COPII coat, a layered protein scaffold that mediates the capture of cargo proteins and the remodeling of the ER membrane into spherical vesicular carriers. Although the components of this machinery have been genetically defined, and the mechanisms of coat assembly extensively explored in vitro, understanding the physical mechanisms of membrane remodeling in cells remains a challenge. Here we use correlative light and electron microscopy (CLEM) to visualize the nanoscale ultrastructure of membrane remodeling at ER exit sites (ERES) in yeast cells.

Refining the definition of the avian pathogenic Escherichia coli (APEC) pathotype through inclusion of high-risk clonal groups.

Colibacillosis in poultry is a unique disease manifestation of Escherichia coli in the animal world, as one of the primary routes of entry is via the respiratory tract of birds. Because of this, a novel extraintestinal pathogenic E. coli (ExPEC) subpathotype coined avian pathogenic E.

The Attachment Versus Differentiation Debate: Bringing the Conversation to Parent-Child Relationships.

This article contributes to the attachment versus differentiation debate, bringing the conversation to parent-child relationships. While attachment theory's (AT) approach emphasizes bonding, Bowen family systems theory's (BFST) differentiation approach emphasizes emotional boundaries. They both suggest that balancing autonomy and connection is important, but AT conceptualizes this in terms of the parent's ability to meet the child's needs for autonomy and connection, while BFST conceptualizes this in terms of the parent's and child's ability to be connected due to mutual respect for each other's emotional boundaries.

Selective inhibition of protein secretion by abrogating receptor-coat interactions during ER export.

Protein secretion is an essential process that drives cell growth, movement, and communication. Protein traffic within the secretory pathway occurs via transport intermediates that bud from one compartment and fuse with a downstream compartment to deliver their contents. Here, we explore the possibility that protein secretion can be selectively inhibited by perturbing protein-protein interactions that drive capture into transport vesicles.

Five Complete Serotype Reading Genomes Recovered from Patients in the United States.

Between 2018 and 2019, Salmonella enterica serotype Reading caused a large, multistate outbreak linked to contact with raw turkey products in the United States. Here, we provide five Salmonella Reading reference genomes collected from US patients between 2016 and 2018.

A SURF4-to-proteoglycan relay mechanism that mediates the sorting and secretion of a tagged variant of sonic hedgehog.

SignificanceSonic Hedgehog (Shh) is a key signaling molecule that plays important roles in embryonic patterning, cell differentiation, and organ development. Although fundamentally important, the molecular mechanisms that regulate secretion of newly synthesized Shh are still unclear. Our study reveals a role for the cargo receptor, SURF4, in facilitating export of Shh from the endoplasmic reticulum (ER) via a ER export signal.

Computed structures of core eukaryotic protein complexes.

Protein-protein interactions play critical roles in biology, but the structures of many eukaryotic protein complexes are unknown, and there are likely many interactions not yet identified. We take advantage of advances in proteome-wide amino acid coevolution analysis and deep-learning–based structure modeling to systematically identify and build accurate models of core eukaryotic protein complexes within the proteome. We use a combination of RoseTTAFold and AlphaFold to screen through paired multiple sequence alignments for 8.

An in vitro vesicle formation assay reveals cargo clients and factors that mediate vesicular trafficking.

The fidelity of protein transport in the secretory pathway relies on the accurate sorting of proteins to their correct destinations. To deepen our understanding of the underlying molecular mechanisms, it is important to develop a robust approach to systematically reveal cargo proteins that depend on specific sorting machinery to be enriched into transport vesicles. Here, we used an in vitro assay that reconstitutes packaging of human cargo proteins into vesicles to quantify cargo capture.