Biology subjects

Hudson, A. W.

Publications and source records attributed to Hudson, A. W..

4 recordsLinked to original sources

bioRxiv2022

Characterization of the HHV-6 U20 immunoevasin

Roseoloviruses (HHV-6A, -6B, and -7) infect >90% of the human population during early childhood, and are thought to remain latent or persistent throughout the life of the host. As such, these viruses are among the most pervasive and stealthy of all viruses; they must necessarily excel at escaping immune detection throughout the life of the host, and yet very little is known about how these viruses so successfully escape host defenses. Herein, we characterize the HHV6A and HHV6B U20 gene products, which are encoded within a block of genes unique to the roseoloviruses, and therefore of particular interest. Despite 92% amino acid identity, U20 proteins from HHV6A and 6B have been shown to possess different host evasion functions. Here we characterize expression, trafficking, and post-translational modifications of U20 during HHV6A infection. While U20 localized to lysosomes in HHV-6A-infected cells, HHV-6B U20 trafficked to the cell surface and was rapidly internalized. HHV-6B U20 trafficked slowly through the secretory system, receiving several post translational modifications to its N-linked glycans indicative of surface expressed glycoproteins. Interestingly, U20 is also phosphorylated on at least one Ser, Thr, or Tyr residue. These results provide a framework to understand the role(s) of U20 in evading host defenses. ImportanceHHV6A and HHV6B U20 proteins are virus-encoded integral membrane glycoproteins possessing class I MHC-like folds. As such, it is tempting to speculate that they are involved in host evasion. Indeed, although they share 92% identity, HHV6A U20 has been shown to target NK activating ligands (1) and HHV6B U20 has been shown to inhibit TNF receptor signaling and apoptosis (2). Here, we have performed cell biological and biochemical characterization of the trafficking, glycosylation, and post-translational modifications occurring on HHV6A and -6B U20, and we demonstrate U20 expression in the context of HHV6 infection.

cell biology↗

bioRxiv2022

Klebsiella pneumoniae L-Fucose metabolism promotes gastrointestinal colonization and modulates its virulence determinants

Colonization of the gastrointestinal (GI) tract by Klebsiella pneumoniae (K. pneumoniae) is generally considered asymptomatic. However, gut colonization allows K. pneumoniae to either translocate to sterile site within the same host or transmit through the fecal-oral route to another host. K. pneumoniae gut colonization is poorly understood, but knowledge of this first step toward infection and spread is critical for combatting its disease manifestations. K. pneumoniae must overcome colonization resistance (CR) provided by the host microbiota to establish itself within the gut. One such mechanism of CR is through nutrient competition. Pathogens that metabolizes a broad range of substrates have the ability to bypass nutrient competition and overcome CR. Herein, we demonstrate that in response to mucin derived fucose, the conserved fucose metabolism operon (fuc) of K. pneumoniae is upregulated in the murine gut and subsequently show that fucose metabolism promotes robust gut colonization. Growth studies using cecal filtrate as a proxy for the gut lumen illustrates the growth advantage that the fuc operon provides K. pneumoniae. We further show that fucose metabolism allows K. pneumoniae to be competitive with a commensal E. coli isolate (Nissle). However, Nissle is eventually able to out-compete K. pneumoniae, suggesting that it can be utilized to enhance CR. Lastly, we observed that fucose metabolism positively modulates hypermucoviscosity, auto-aggregation, and biofilm formation, but not capsule biogenesis. Together, these insights enhance our understanding of the role of alternative carbon sources on K. pneumoniae gut colonization and the complex relationship between metabolism and virulence in this species. ImportanceKlebsiella pneumoniae (K. pneumoniae) is a leading cause of hospital-acquired infection. Treatment of infection by K. pneumoniae isolates is becoming difficult as this pathogen becomes increasingly antibiotic resistant. While there has been extensive investigation into the disease states associated with K. pneumoniae, its colonization of the gastro-intestinal (GI) tract is poorly understood. Epidemiological data suggest that in many cases the strain that colonizes the GI tract causes disease manifestations in the same host. Herein, we used our newly developed murine model of K. pneumoniae gut colonization, where colonization is achieved without disrupting the resident gut microbiota. We demonstrate that K. pneumoniae uses fucose as an alternative carbon source present in the gut lumen to overcome the intense nutritional competition. We further illustrate that K. pneumoniae, through fucose metabolism, is initially competitive with the probiotic E coli isolate Nissle 1917 (EcN). Lastly, we show that fucose metabolism modulates several virulence determinants of K. pneumoniae. Thus, our results provide new insight into the role fucose metabolism plays in gut colonization and virulence of K. pneumoniae, and furthermore identify EcN as having the ability to out-compete K. pneumoniae and be used as a probiotic.

microbiology↗

bioRxiv2021

MgrB dependent colistin resistance in Klebsiella pneumoniae is associated with an increase in host-to-host transmission.

Due to its high transmissibility, Klebsiella pneumoniae (Kpn) is one of the leading causes of nosocomial infections. Here, we studied the biological cost of colistin resistance, an antibiotic of last resort, of this opportunistic pathogen using a murine model of gut colonization and transmission. Colistin resistance in Kpn is commonly the result of inactivation of the small regulatory protein MgrB. Without a functional MgrB, the two-component system PhoPQ is constitutively active, leading to increased lipid A modifications and subsequent colistin resistance. Using an engineered MgrB mutant, we observed that MgrB-dependent colistin resistance is not associated with a fitness defect during in vitro growth conditions. However, colistin-resistant Kpn colonizes the murine gut poorly, which may be due to the decreased production of capsular polysaccharide by the mutant. The colistin-resistant mutant of Kpn had increased survival outside the host when compared to the parental colistin-sensitive strain. We attribute this enhanced survivability to dysregulation of the PhoPQ two-component system and accumulation of the master stress regulator RpoS. The enhanced survival of the colistin resistant strain may be a key factor in the observed rapid host-to-host transmission in our model. Together, our data demonstrate that colistin-resistant Kpn experiences a biological cost in gastrointestinal colonization. However, this cost is mitigated by enhanced survival outside the host, increasing the risk of transmission. Additionally, it underscores the importance of considering the entire life cycle of a pathogen to truly determine the biological cost associated with antibiotic resistance. ImportanceThe biological cost associated with colistin resistance in Klebsiella pneumoniae (Kpn) was examined using a murine model of Kpn gut colonization and fecal-oral transmission. A common mutation resulting in colistin resistance in Kpn is a loss-of-function mutation of the small regulatory protein MgrB that regulates the two-component system PhoPQ. Even though colistin resistance in Kpn comes with a fitness defect in gut colonization, it increases bacterial survival outside the host enabling it to more effectively transmit to a new host. The enhanced survival is dependent upon the accumulation of RpoS and dysregulation of the PhoPQ. Hence, our study expands our understanding of the underlying molecular mechanism contributing to the transmission of colistin-resistant Kpn.

microbiology↗

bioRxiv2020

The roseoloviruses downregulate the protein tyrosine phosphatase PTPRC(CD45)

Like all herpesviruses, the roseoloviruses (HHV6A, -6B, and -7) establish lifelong infection within their host, requiring these viruses to evade host anti-viral responses. One common host-evasion strategy is the downregulation of host-encoded, surface-expressed glycoproteins. Roseoloviruses have been shown to evade host the host immune response by downregulating NK-activating ligands, MHC class I, and the TCR/CD3 complex. To more globally identify glycoproteins that are differentially expressed on the surface of HHV6A-infected cells, we performed cell surface capture of N-linked glycoproteins present on the surface of T cells infected with HHV6A, and compared these to proteins present on the surface of uninfected T cells. We found that the protein tyrosine phosphatase CD45 is downregulated in T cells infected with HHV6A. We also demonstrated that CD45 is similarly downregulated in cells infected with HHV-7. CD45 is essential for signaling through the T cell receptor and as such, is necessary for developing a fully functional immune response. Interestingly, the closely related {beta}-herpesviruses human cytomegalovirus (HCMV) and murine cytomegalovirus (MCMV) have also separately evolved unique mechanisms to target CD45. While HCMV and MCMV target CD45 signaling and trafficking, HHV6A acts to downregulate CD45 transcripts. ImportanceHuman herpesviruses-6 and -7 infect essentially 100% of the worlds population before the age of 5 and then remain latent or persistent in their host throughout life. As such, these viruses are among the most pervasive and stealthy of all viruses. Host immune cells rely on the presence of surface-expressed proteins to identify and target virus-infected cells. Here, we investigated the changes that occur to proteins expressed on the cell surface of T cells after infection with human herpesvirus-6A. We discovered that HHV-6A infection results in a reduction of CD45 on the surface of infected cells. Targeting of CD45 may prevent activation of these virus-infected T cells, possibly lengthening the life of the infected T cell so that it can harbor latent virus.

microbiology↗