Tubulin acetyltransferases access and modify the microtubule luminal K40 residue through anchors in taxane-binding pockets.

Acetylation at α-tubulin K40 is the sole post-translational modification preferred to occur inside the lumen of hollow cylindrical microtubules. However, how tubulin acetyltransferases access the luminal K40 in micrometer-long microtubules remains unknown. Here, we use cryo-electron microscopy and single-molecule reconstitution assays to reveal the enzymatic mechanism for tubulin acetyltransferases to modify K40 in the lumen.

Replication licensing regulated by a short linear motif within an intrinsically disordered region of origin recognition complex.

In eukaryotes, the origin recognition complex (ORC) faciliates the assembly of pre-replicative complex (pre-RC) at origin DNA for replication licensing. Here we show that the N-terminal intrinsically disordered region (IDR) of the yeast Orc2 subunit is crucial for this process. Removing a segment (residues 176-200) from Orc2-IDR or mutating a key isoleucine (194) significantly inhibits replication initiation across the genome.

Assembly and activation of replicative helicases at origin DNA for replication initiation.

To initiate DNA replication, it is essential to properly assemble a pair of replicative helicases at each replication origin. While the general principle of this process applies universally from prokaryotes to eukaryotes, the specific mechanisms governing origin selection, helicase loading, and subsequent helicase activation vary significantly across different species. Recent advancements in cryo-electron microscopy (cryo-EM) have revolutionized our ability to visualize large protein or protein-DNA complexes involved in the initiation of DNA replication.

Synergism between CMG helicase and leading strand DNA polymerase at replication fork.

The replisome that replicates the eukaryotic genome consists of at least three engines: the Cdc45-MCM-GINS (CMG) helicase that separates duplex DNA at the replication fork and two DNA polymerases, one on each strand, that replicate the unwound DNA. Here, we determined a series of cryo-electron microscopy structures of a yeast replisome comprising CMG, leading-strand polymerase Polε and three accessory factors on a forked DNA. In these structures, Polε engages or disengages with the motor domains of the CMG by occupying two alternative positions, which closely correlate with the rotational movement of the single-stranded DNA around the MCM pore.

Menin "reads" H3K79me2 mark in a nucleosomal context.

Methylation of histone H3 lysine-79 (H3K79) is an epigenetic mark for gene regulation in development, cellular differentiation, and disease progression. However, how this histone mark is translated into downstream effects remains poorly understood owing to a lack of knowledge about its readers. We developed a nucleosome-based photoaffinity probe to capture proteins that recognize H3K79 dimethylation (H3K79me2) in a nucleosomal context.

The human pre-replication complex is an open complex.

In eukaryotes, DNA replication initiation requires assembly and activation of the minichromosome maintenance (MCM) 2-7 double hexamer (DH) to melt origin DNA strands. However, the mechanism for this initial melting is unknown. Here, we report a 2.

Association between initial symptoms and subsequent hospitalization in outpatients with COVID-19: A cohort study.

Background: Most individuals with coronavirus disease 2019 (COVID-19) experience mild symptoms and are managed in the outpatient setting. The aim of this study was to determine whether self-reported symptoms at the time of diagnosis can identify patients at risk of clinical deterioration.

Methods: This was a retrospective cohort study of 671 outpatients with laboratory-confirmed COVID-19 diagnosed in Toronto between March 1 and October 16, 2020.

Humanizing the yeast origin recognition complex.

The Origin Recognition Complex (ORC) is an evolutionarily conserved six-subunit protein complex that binds specific sites at many locations to coordinately replicate the entire eukaryote genome. Though highly conserved in structure, ORC's selectivity for replication origins has diverged tremendously between yeasts and humans to adapt to vastly different life cycles. In this work, we demonstrate that the selectivity determinant of ORC for DNA binding lies in a 19-amino acid insertion helix in the Orc4 subunit, which is present in yeast but absent in human.

Mp1 Protein, a Novel Virulence Factor, Carries Two Arachidonic Acid-Binding Domains To Suppress Inflammatory Responses in Hosts.

infection causes talaromycosis (previously known as penicilliosis), a very important opportunistic systematic mycosis in immunocompromised patients. Different virulence mechanisms in have been proposed and investigated. In the sera of patients with talaromycosis, Mp1 protein (Mp1p), a secretory galactomannoprotein antigen with two tandem ligand-binding domains (Mp1p-LBD1 and Mp1p-LBD2), was found to be abundant.

Structure of the origin recognition complex bound to DNA replication origin.

The six-subunit origin recognition complex (ORC) binds to DNA to mark the site for the initiation of replication in eukaryotes. Here we report a 3 Å cryo-electron microscopy structure of the Saccharomyces cerevisiae ORC bound to a 72-base-pair origin DNA sequence that contains the ARS consensus sequence (ACS) and the B1 element. The ORC encircles DNA through extensive interactions with both phosphate backbone and bases, and bends DNA at the ACS and B1 sites.

Talaromyces marneffei Mp1p Is a Virulence Factor that Binds and Sequesters a Key Proinflammatory Lipid to Dampen Host Innate Immune Response.

Talaromyces (Penicillium) marneffei is one of the leading causes of systemic mycosis in immunosuppressed or AIDS patients in Southeast Asia. How this intracellular pathogen evades the host immune defense remains unclear. We provide evidence that T.

Mp1p Is a Virulence Factor in Talaromyces (Penicillium) marneffei.

Background: Talaromyces marneffei is an opportunistic dimorphic fungus prevalent in Southeast Asia. We previously demonstrated that Mp1p is an immunogenic surface and secretory mannoprotein of T. marneffei.