The proteasome regulator PSME4 modulates proteasome activity and antigen diversity to abrogate antitumor immunity in NSCLC.

Immunotherapy revolutionized treatment options in cancer, yet the mechanisms underlying resistance in many patients remain poorly understood. Cellular proteasomes have been implicated in modulating antitumor immunity by regulating antigen processing, antigen presentation, inflammatory signaling and immune cell activation. However, whether and how proteasome complex heterogeneity may affect tumor progression and the response to immunotherapy has not been systematically examined. Here, we show that proteasome complex composition varies substantially across cancers and impacts tumor-immune interactions and the tumor microenvironment. Through profiling of the degradation landscape of patient-derived non-small-cell lung carcinoma samples, we find that the proteasome regulator PSME4 is upregulated in tumors, alters proteasome activity, attenuates presented antigenic diversity and associates with lack of response to immunotherapy. Collectively, our approach affords a paradigm by which proteasome composition heterogeneity and function should be examined across cancer types and targeted in the context of precision oncology.

Crystal structure of the MrkD1P receptor binding domain of Klebsiella pneumoniae and identification of the human collagen V binding interface.

Klebsiella species are members of the family enterobacteriaceae, opportunistic pathogens that are among the eight most prevalent infectious agents in hospitals. Among other virulence factors in Klebsiella, type 3 pili exhibit a unique binding pattern in the human kidney via interaction of two MrkD adhesion variants 1C1 and 1P to type IV and/or V collagen. However, very little is known about the nature of this recognition. Here we present the crystal structure of the plasmid born MrkD1P receptor domain (MrkDrd). The structure reveals a jelly-roll ß-barrel fold comprising 17 ß-strands very similar to the receptor domain of GafD, the tip adhesin from the F17 pilus that recognizes n-acetyl-d-glucosamine (GlcNAc). Analysis of collagen V binding of different MrkD1P mutants revealed that two regions were responsible for its binding: a pocket, that aligns approximately with the GlcNAc binding pocket of GafD involving residues R105 and Y155, and a transversally oriented patch that spans strands ß2a, ß9b and ß6 including residues V49, T52, V91, R102 and I136. Taken together, these data provide structural and functional insights on MrkD1P recognition of host cells, providing a tool for future development of rationally designed drugs with the prospect of blocking Klebsiella adhesion to collagen V.

Structural homology between the C-terminal domain of the PapC usher and its plug.

P pili are extracellular appendages responsible for the targeting of uropathogenic Escherichia coli to the kidney. They are assembled by the chaperone-usher (CU) pathway of pilus biogenesis involving two proteins, the periplasmic chaperone PapD and the outer membrane assembly platform, PapC. Many aspects of the structural biology of the Pap CU pathway have been elucidated, except for the C-terminal domain of the PapC usher, the structure of which is unknown. In this report, we identify a stable and folded fragment of the C-terminal region of the PapC usher and determine its structure using both X-ray crystallography and nuclear magnetic resonance (NMR) spectroscopy. These structures reveal a beta-sandwich fold very similar to that of the plug domain, a domain of PapC obstructing its translocation domain. This structural similarity suggests similar functions in usher-mediated pilus biogenesis, playing out at different stages of the process. This structure paves the way for further functional analysis targeting surfaces common to both the plug and the C-terminal domain of PapC.

Two-step and one-step secretion mechanisms in Gram-negative bacteria: contrasting the type IV secretion system and the chaperone-usher pathway of pilus biogenesis.

Gram-negative bacteria have evolved diverse secretion systems/machineries to translocate substrates across the cell envelope. These various machineries fulfil a wide variety of functions but are also essential for pathogenic bacteria to infect human or plant cells. Secretion systems, of which there are seven, utilize one of two secretion mechanisms: (i) the one-step mechanism, whereby substrates are translocated directly from the bacterial cytoplasm to the extracellular medium or into the eukaryotic target cell; (ii) the two-step mechanism, whereby substrates are first translocated across the bacterial inner membrane; once in the periplasm, substrates are targeted to one of the secretion systems that mediate transport across the outer membrane and released outside the bacterial cell. The present review provides an example for each of these two classes of secretion systems and contrasts the various solutions evolved to secrete substrates.

1,3-Propanediol dehydrogenase from Klebsiella pneumoniae: decameric quaternary structure and possible subunit cooperativity.

Klebsiella pneumoniae is a nosocomial pathogen frequently isolated from opportunistic infections, especially in clinical environments. In spite of its potential pathogenicity, this microorganism has several metabolic potentials that could be used in biotechnology applications. K. pneumoniae is able to metabolize glycerol as a sole source of carbon and energy. 1,3-Propanediol dehydrogenase is the core of the metabolic pathway for the use of glycerol. We have determined the crystallographic structure of 1,3-propanediol dehydrogenase, a type III Fe-NAD-dependent alcohol dehydrogenase, at 2.7-A resolution. The structure of the enzyme monomer is closely related to that of other alcohol dehydrogenases. The overall arrangement of the enzyme showed a decameric structure, formed by a pentamer of dimers, which is the catalytic form of the enzyme. Dimers are associated by strong ionic interactions that are responsible for the highly stable in vivo packing of the enzyme. Kinetic properties of the enzyme as determined in the article would suggest that this decameric arrangement is related to the cooperativity between monomers.

CYP21A2 mutations in Portuguese patients with congenital adrenal hyperplasia: identification of two novel mutations and characterization of four different partial gene conversions.

More than 90% of congenital adrenal hyperplasia (CAH) cases are caused by 21-hydroxylase deficiency. In this study, the CYP21 gene was genotyped in 56 Portuguese unrelated patients with clinical symptoms of 21-hydroxylase deficiency, in a total of 112 independent alleles. CYP21A2 mutations were identified in 99.1% of the alleles. The most common point mutation was 1688G>T (25.9%). A previously unreported partial gene conversion, extending from exon 1 to 7, was found in 16.1% of the alleles, in most cases associated to the mutation 1688G>T in the other chromosome, and in patients with nonclassical CAH. Other three distinct partial gene conversions were also identified, with lower frequencies: one extends from exon 1 to 3 and the others from exons 3 to 7 and 3 to 8. Two novel mutations were identified in two salt-wasting patients: a putative splicing mutation, IVS2+5G>A, and the transition 2557C>T, that gives rise to the nonsense mutation R445X. Seven point mutations and a partial gene conversion were responsible for 88 of the studied disease causing alleles, and the overall concordance between genotype and phenotype was 92.9%. With this study the molecular basis of CAH was characterized, for the first time, in Portuguese patients, providing useful results for clinicians in terms of prediction of disease severity, genetic and prenatal counseling.