Patients who survive sepsis display suppressed immune functions, often manifested as an increased susceptibility to secondary infections. Recently, using a cecal-ligation and puncture (CLP) model of sepsis we showed that sepsis induces substantial and long-lasting changes in the available naive CD8+ T cell repertoire affecting the capacity of the host to respond to newly encountered acute infections. However, the extent to which sepsis changes the host susceptibility to chronic infection and affects CD8+ T cell responses is currently unknown. Here, we demonstrate that inbred and outbred mice recovering from a septic event are more susceptible to LCMV clone-13 infection exhibited by mortality and viral burden. Primary virus-specific CD8+ T cells in LCMV clone-13 infected septic mice displayed exacerbated CD8+ T cell exhaustion illustrated by increased inhibitory molecule expression (e.g., PD-1, LAG-3, and 2B4) and diminished Ag-driven cytokine production (e.g. IFNγ, TNFα) compared to similarly infected sham-treated mice. Importantly, therapeutic inhibitory molecule dual-blockade (αPD-L1 and αLAG-3) increased the number of circulating LCMV-specific CD8+ T cells, improved CD8+ T cell function and pathogen control in chronically infected septic mice. Together, these results illustrate that poly-microbial sepsis compromises the overall health of the host leading to increased vulnerability to chronic infection and exacerbated CD8+ T cell exhaustion. Collectively, our findings suggest that septic survivors may be more susceptible and at higher risk of developing exhaustible CD8+ T cells upon encountering a subsequent chronic infection.

In nonneuronal cells, herpes simplex virus 1 overcomes host defenses, replicates, and ultimately kills the infected cell. Among the host defenses suppressed by the virus is a repressor complex whose key components are histone deacetylase (HDAC) 1 or 2, RE-1 silencing transcription factor (REST), corepressor of REST (CoREST), and lysinespecific demethylase (LSD) 1. In neurons innervating cells at the portal of entry into the body, the virus establishes a "latent" infection in which viral DNA is silenced with the exception of a family of genes. The question posed here is whether the virus hijacks this repressor complex to silence itself in neurons during the latent state. To test this hypothesis, we inserted into the wild-type virus genome a wild-type REST [recombinant (R) 111], a dominant-negative REST (dnREST) lacking the N-and C-terminal repressor domains (R112), or an insertion control consisting of tandem repeats of stop codons (R113). The recombinant virus R112 carrying the dnREST replicated better and was more virulent than the wild-type parent or the other recombinant viruses when administered by the corneal or i.p. routes. Moreover, in contrast to other recombinants, corneal route inoculation by R112 recombinant virus resulted in higher DNA copy numbers, higher levels of infectious virus in eye, trigeminal ganglion, or brain, and virtually complete destruction of trigeminal ganglia in mice that may ultimately succumb to infection. These results support an earlier conclusion that the HDAC/CoREST/REST/LSD1 repressor complex is a significant component of the host innate immunity and are consistent with the hypothesis that HSV-1 hijacks the repressor to silence itself during latent infection.chromatin remodeling | herpesviruses | latency H erpes simplex virus 1 (HSV-1) replicates at the portal of entry into the body, infects sensory nerve endings, and is transported retrograde to the neuronal nucleus (reviewed in ref. 1). In mice, a common animal model system, the virus replicates in some neurons but is silenced and establishes a latent infection in other neurons. Thus, infectious virus is readily detected during the first 10-15 d after infection at a peripheral site. It then disappears, and by day 28 only latent virus is present in the ganglia. One explanation for the two different outcomes of infection is that on release of the viral DNA into the nucleus, the cell attempts to silence the DNA. This attempt leads to a silent, latent infection in neurons but not in cells at the portal of entry into the body or in cell cultures in which the virus replicates. The fundamental question posed in the studies reported here is whether the silencing system suppressed by the virus in productively infected cells is enabled to establish a silent, "latent" infection in neurons.Specifically, several lines of evidence indicate that in infected cells, ICP0, an α (immediate-early) protein, plays a crucial role in enabling viral replication at low multiplicities of infection. At the portal of entry, the genes encoding α protein...

Abstractβ-Coronaviruses such as SARS-CoV-2 hijack coatomer protein-I (COPI) for spike protein retrograde trafficking to the progeny assembly site in endoplasmic reticulum-Golgi intermediate compartment (ERGIC). However, limited residue-level details are available into how the spike interacts with COPI. Here we identify an extended COPI binding motif in the spike that encompasses the canonical K-x-H dibasic sequence. This motif demonstrates selectivity for αCOPI subunit. Guided by an in silico analysis of dibasic motifs in the human proteome, we employ mutagenesis and binding assays to show that the spike motif terminal residues are critical modulators of complex dissociation, which is essential for spike release in ERGIC. αCOPI residues critical for spike motif binding are elucidated by mutagenesis and crystallography and found to be conserved in the zoonotic reservoirs, bats, pangolins, camels, and in humans. Collectively, our investigation on the spike motif identifies key COPI binding determinants with implications for retrograde trafficking.

Memory CD8 T cells play a critical role in providing protection to immune hosts by orchestrating rapid elimination of pathogen-infected cells after re-infection. Systemic bacterial infection with Listeria monocytogenes has been a favored approach for researchers to characterize pathogen-specific CD8 T cell responses, and in-depth understanding of L. monocytogenes biology has provided invaluable experimental tools that have been used to increase our understanding of memory CD8 T cell differentiation. Here, we describe how the tools from this murine model system of infection have been utilized to characterize pathogen-specific CD8 T cells in inbred and genetically diverse outbred hosts as they undergo naïve-to-memory CD8 T cell differentiation in vivo. We also discuss how studying L. monocytogenes-evoked CD8 T cell responses have provided insight on the degree of diminished T cell immunity in clinically relevant conditions such as sepsis and obesity. Overall, this review will highlight how infection with the intracellular pathogen L. monocytogenes has enabled analysis of systemic CD8 T cell responses and greatly contributed to what is known about memory CD8 T cell generation and differentiation.

The extent to which obesity compromises the differentiation and maintenance of protective memory CD8 T cell responses and renders obese individuals susceptible to infection remains unknown. Here, we show that diet-induced obesity did not impact the maintenance of pre-existing memory CD8 T cells including acquisition of a long-term memory phenotype (i.e. CD27hi, CD62Lhi, KLRG1low) and function (i.e. cytokine production, secondary expansion, and memory CD8 T cell-mediated protection). In addition, obesity did not influence the differentiation and maintenance of newly evoked memory CD8 T cell responses in inbred and outbred hosts generated in response to different types of systemic (LCMV, L. monocytogenes) and/or localized (influenza virus) infections. Interestingly, the rate of naïve-to-memory CD8 T cell differentiation after a peptide-coated dendritic cell (DC) immunization was similar in lean and obese hosts suggesting that obesity associated inflammation, unlike pathogen- or adjuvant-induced inflammation, did not influence the development of endogenous memory CD8 T cell responses. Therefore, our studies reveal that the obese environment does not influence the development or maintenance of memory CD8 T cell responses that are either primed before or after obesity is established, a surprising notion with important implications for future studies aiming to elucidate the role obesity plays in host susceptibility to infections.

Highlights d The structure of the human calpain-5 protease core (CAPN5-PC) is determined d CAPN5-PC contains three elongated loops compared to its classical counterparts d One loop contains a mutation identified in a patient with NIV (p.Gly267Ser) d The p.Gly267Ser mutation causes hyperactivity in CAPN5-PC and CAPN1/5-PC hybrids

Calpain-5 is a calcium-activated protease expressed in the retina. Mutations in calpain-5 cause autosomal dominant neovascular inflammatory vitreoretinopathy (ADNIV, OMIM#193235). The structure of calpain-5 has not been determined, thus hindering the investigation of its proteolytic targets and pathological role in ADNIV. Herein, we report models of the proteolytic core of calpain-5 (mini-calpain-5) containing two globular domains (termed DIIa-IIb) connected by a short, flexible linker, consistent with small-angle x-ray scattering (SAXS) data. Structural modeling in the absence of calcium suggests that mini-calpain-5 adopts a more open conformation when compared to previously determined structures of other calpain cores. This open conformation, achieved by a rotation of DIIa and DIIb with respect to each other, prevents formation of the active site and constrains the enzyme in an inactivated form. The relative domain rotation of 60-100°we found for mini-calpain-5 (a non-classical calpain) is significantly greater than the largest rotation previously observed for a classical calpain (i.e., 55.0°for mini-calpain-9). Together with our prediction that, in the full-length form, a long loop in DIIb (loop C1), a few residues downstream of the inter-domain linker, likely interacts with the shorter, acidic, inactivating loop on domain-III (DIII), these structural insights illuminate the complexity of calpain regulation. Moreover, our studies argue that pursuing higher resolution structural studies are necessary to understand the complex activity regulation prevalent in the calpain family and for the design of specific calpain inhibitors.

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