1.
Enhanced ER proteostasis and temperature differentially impact the mutational tolerance of influenza hemagglutinin.
Phillips, AM, Doud, MB, Gonzalez, LO, Butty, VL, Lin, YS, Bloom, JD, Shoulders, MD
eLife. 2018
Abstract
We systematically and quantitatively evaluate whether endoplasmic reticulum (ER) proteostasis factors impact the mutational tolerance of secretory pathway proteins. We focus on influenza hemaggluttinin (HA), a viral membrane protein that folds in the host's ER via a complex pathway. By integrating chemical methods to modulate ER proteostasis with deep mutational scanning to assess mutational tolerance, we discover that upregulation of ER proteostasis factors broadly enhances HA mutational tolerance across diverse structural elements. Remarkably, this proteostasis network-enhanced mutational tolerance occurs at the same sites where mutational tolerance is most reduced by propagation at fever-like temperature. These findings have important implications for influenza evolution, because influenza immune escape is contingent on HA possessing sufficient mutational tolerance to evade antibodies while maintaining the capacity to fold and function. More broadly, this work provides the first experimental evidence that ER proteostasis mechanisms define the mutational tolerance and, therefore, the evolution of secretory pathway proteins.
2.
Ca²+ signaling in B cells.
Lyubchenko, T
TheScientificWorldJournal. 2010;:2254-64
Abstract
An increase in intracellular Ca²+ concentration is one of the major initial steps in B-cell activation that occurs within minutes after antigen receptor (BCR) engagement. In recent years, significant advances have been made in characterizing molecular mechanisms of Ca²+ signaling in lymphocytes, although the majority of work was done on T cells. This mini-review discusses several underexplored areas of Ca²+ signaling in B cells: (1) Ca²+ signaling in immune synapse and multifaceted Ca²+ responses within a single cell, (2) source of Ca²+ involved in Ca²+-dependent protein phosphorylation events and the role of store-operated influx, (3) role of BCR coreceptors in Ca²+ signaling, and (4) Ca²+ signaling and maintenance of B-cell tolerance and clinical significance of Ca²+ signaling alterations.
3.
The adenovirus E3-6.7K protein adopts diverse membrane topologies following posttranslational translocation.
Moise, AR, Grant, JR, Lippé, R, Gabathuler, R, Jefferies, WA
Journal of virology. 2004;(1):454-63
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Abstract
The E3 region of adenovirus codes for several membrane proteins, most of which are involved in immune evasion and prevention of host cell apoptosis. We explored the topology and targeting mechanisms of E3-6.7K, the most recently described member of this group, by using an in vitro translation system supplemented with microsomes. Here, we present evidence that E3-6.7K, one of the smallest signal-anchor proteins known, translocates across the membrane of the endoplasmic reticulum in a posttranslational, ribosome-independent, yet ATP-dependent manner, reminiscent of the translocation of tail-anchored proteins. Our analysis also demonstrated that E3-6.7K could achieve several distinct topological fates. In addition to the previously postulated type III orientation (N-luminal/C-cytoplasmic, termed NtmE3-6.7K), we detected a tail-anchored form adopting the opposite orientation (N-cytoplasmic/C-luminal, termed CtmE3-6.7K) as well as the possibility of a fully translocated form (N and C termini are both translocated, termed NCE3-6.7K). Due to the translocation of a positively charged domain, both the CtmE3-6.7K and NCE3-6.7K topologies of E3-6.7K constitute exceptions to the "positive inside" rule. The NtmE3-6.7K and NCE3-6.7K are the first examples of posttranslationally translocated proteins in higher eukaryotes that are not tail anchored. Distinct topological forms were also found in transfected cells, as both N and C termini of E3-6.7K were detected on the extracellular surface of transfected cells. The demonstration of unexpected topological forms and translocation mechanisms for E3-6.7K defies conventional thinking about membrane protein topogenesis and advises that both the mode of targeting and topology of signal-anchor proteins should be determined experimentally.