Stimulatory cytokines for ILC-2s
IL-33 (strong response) and IL-25 (moderate response)
How does T2 immunity lead to helminth expulsion?
In addition to signals that activate the `epithelial escalator’, the type 2 cytokines IL-4, IL-5, IL-9 and IL-13 contribute to increased muscle contractility, diarrhea and changes to the mucus that lines the gastrointestinal tract
What are the steps of tissue repair after damage in epithelial layer?
Following cellular stress or tissue damage, adenosine is released at the site of inflammation, which can induce up-regulation of the alarmin IL-33 and enhance the development of tissue- protective M2 macrophages and eosinophils at the site of parasite infection. IL-33 activates ILC2s which release amphiregulin.
What is a role for amphiregulin?
Amphiregulin from ILC2s leads to tissue repair and protection from DSS-induced damage as well as damage from influenza infection.
thermogenic protein in beige adipose tissue that is critical for burning calories and preventing obesity.
What is a phenotype of ILC2- or IL-33-deficient mice?
Reduced beige adipose, increased obesity. Loss of tissue repair mechanisms.
How do ILC2s regulate adipose beiging?
ILC2-derived enkephalin peptides promote beiging and tissue protection of white adipose. This leads to an increase in energy expenditure and a decrease in adiposity.
What cytokines activate ILC2s and which cytokines do they produce in response?
ILC2s are activated by IL-25, IL-33, and TSLP, and produce Amphiregulin, IL-13, IL-5, and IL-9,
What is the effect of ILC2 cytokines on other cells?
Epithelial cells - mucus production “weep and sweep”
Eosinophil activation and production of cytokines
Mast cell activation
inflammatory ILC2s phwnotype
IL-33 required for their generation
Tryptophan hydroxylase (Tph)
rate-limiting enzyme of the biosynthesis of serotonin (5-HT) from its precursor L-tryptophan
2 isoforms of Tph
Tph1 - Non-neuronal cells: Enterochromaffin cells and mast cells (and ILC2)
Tph2 - Neurons: brainstem and myenteric plexus
Which mRNAs are upregulated in ILC2 after IL-33 treatment?
mki67 il5 il13 Areg Tph1
What is the phenotype of mice lacking Tph1 in lymphocytes?
reduced inflammatory ILC2s, reduced worm expulsion in helminth infections.
ILC2 main transcription factor
Neuromedin U (Nmur1) is selectively expressed by ILC2s and not expressed by eosinophils, mast cells, basophil or other hematopoietic cells.
ILC2s co-localize with NMU-expressing neurons in the intestinal submucosa.
Stimulation of Nmur1 on ILC2s leads to production of IL13 and IL5
What cells do ILC2 co localize with in the intestinal submucosa?
NMU-positive cholinergic neurons
Role of NMU in the gut
directly stimulates ILC2s,
promotes type 2 responses and accelerates worm expulsion and type 2 inflammation in the lung.
Beta-2 adrenergic receptors in ILC2ss
Relatively upregulated expression on ILC2s. ILC2s are closely colocalized with neurons that produce the ligand for this receptor (norepinephrine) , and receptor KO mice have enhanced ILC2 responses, suggesting that B2 adrenergic receptor signaling may suppress ILC2s
What might be the neuronal ILC2 ‘rheostat’?
Cholengeric neurons activate ILC2s through NMU/Nmur1 signaling, and Catecholaminergic neurons inhibit ILC2s through norepinephrine/B2AR signaling.
enzyme involved in cell wall (septum) synthesis in E. coli
Five tasks that bacteria must content with when exporting proteins
discriminate the cytoplasmic-resident proteins from those that are destined for export;
deal with the inherent tendency of polypeptides to fold rapidly;
target exported proteins to the membrane with specificity and fidelity;
achieve transmembrane crossing of these elongated, heteropolymeric substrates, which are several times as long as the membrane is wide; and finally,
manage a second sorting event that releases membrane proteins into the lipid bilayer and secretory proteins to the trans side of the membrane
Which secretion pathway is ubiquitous and essential for all three domains of life?
the Sec pathway, which additionally acts as the entry point for many of the other protein export and sorting pathways
What are the two pathways for protein targeting for secretion via the Sec pathway?
SRP-mediated targeting (co-trasnlational)
SecB-mediated targeted (co- and post-translational)
What structure is the T3SS related to?
Cysteine protease, prevents activation of MAPK and NFkB signalling
Yesrinia T3SS protein, inhibits Rho GTPases (disruption of actin filaments)
Yersinia T3SS protein, a tyrosine phosphatase, which dephosphorylates key actin cytoskeletal proteins and inhibits Ca2+ signaling.
Different pathogenicity islands of Salmonella and their functions
SPI-1: epithelial cell invasion/apoptosis
SPI-2, -3, and -4: intramacrophage survival
What is the difference between primary pathogens and opportunistic pathogens?
- Primary pathogens are capable of causing overt disease in healthy (immuncompetent) hosts
- Opportunistic pathogens primarily cause disease in immunocompromised hosts
virulence factor produced bacteria that force non-phagocytic cells to engulf bacteria
Which virulence factors allow for bacteria to survive within phagosomes?
Catalases and superoxide dismutase can reduce and resist oxidative burst
Which motifs are required for surface avaialbility of CpnT in Mtb?
ESX motifs allow for the targeting of mycobacterial proteins to the Esx secretion system. In CpnT, both “Y1” and “Y3” are required for insertion of CpnT into the outer membrane of Mtb.
Which secretion system is essential for the surface avaialbility of CpnT in Mtb?
The ESX-4 system, encoded by the EccC4 gene. ESX-3 and ESX-5 seem to be dispensable.
Also important is the ESXF and ESXE complex, which also erequires ESX-4 system.
How did authors determine whether TNT was intraphagosomal or cytosolic?
By using different permeabilization methods:
Digitonin permeabilizes only the plasma membrane of host cells, whereas Triton X-100 will permeabilize both the cell membrane and the phagosome.
How is the Mtb toxin TNT cytotoxic?
Tuberculosis Necrotizing Toxin (TNT) gains access to the cytosol of infected macrophages and depletes NAD+, it is a novel NAD+ glycohydrolase. When NAD+ levels drops below a certain threshold, RIPK3 is activated and programmed cell death occurs.
What are the steps for TNT export in MtB?
Within the macrophage phagosome, Mtb first makes esxE/esxF, which is exported through ESX-4 into the periplasmic space and inserted into the outer membrane as a pore channel. CpnT is produced and targeted to ESX-4 secretion through the Y1 and Y3 ESX motifs, and is subsequently inserted into the outer membrane by the esxEF pore complex. Finally, TNT is cleaved from the cell surface CpnT by an unknown mechanisms.
Host protein that recognizes luminal glycans that become exposed to the cytosol when a phagosome is extensively damaged. Used to monitor phagosomal disruption in Mtb infection models.
Which proteins allow for Mtb access to the cytosol from the phagosome?
EsxA:EsxB heterodimer secreted by the ESX-1 system is separated into componet parts and EsxA is a membranolytic protein that disrupts the phagosomal membrane.
PDIM in Mtb virulence
Phthiocerol dimycoserosate (PDIM) is a lipid virulence factor of Mtb that allows for phagosomal escape by Mtb. PDIM can potentiate EsxA membranolytic activity by either synergizing the pore forming activity of EsxA or by acting on the EsxA/B heterodimer complex separation
What are some intraphagosomal stressors for Mtb that it has evolved genetic solutions for?
Reactive oxygen species, Hypoxia, Starvation (amino acid, nutrient, etc), low pH, Nutritional Immunity (Cu, Zn)
Site-2 proteases (S2Ps)
widely distributed in bacteria and participate in diverse pathways, all of which share the requirement for proteolysis of a transmembrane protein
How does proteolytic signal transduction work in many bacteria?
A site-1 protease (S1P) first cleaves the (usually) extracytoplasmic segment of the transmembrane substrate in response to specific inducing signal.
This site-1 cleavage is rapidly followed by S2P cleavage within the transmembrane segment of the substrate, thereby liberating the cytosolic fragment of the substrate (often a sigma factor).
Rip1 in Mtb
Proteolytic signal transduction machinery in Mtb that is essetial for the localization for principal mycolic acids into the outer envelope and thus Mtb is attenuated in its absence.
Why is it difficult to determine the underlying mechanisms of resistance in Mtb infections?
All reistance in TB is chromosomally acquired, yet we do not understand the mechanisms of mutagenesis in mycobacteria.
What is required for the IFNg-independent control of Mtb infection?
CD4 T cell production of GM-CSF and activation of the macrophage transcription factor HIFa, which drives a metabolic shift towards aerobic glycolysis and leads to the production of lipid droplets.
Why has the central dogma of IFNg-producing CD4 T cells come under increasing scrutiny as the sole immunological process through which protection to Mtb infection is conferred?
the frequency of IFNg-secreting CD4 T cells correlates more closely with disease severity than with protection from TB disease after M. tuberculosis challenge
human trials with the vaccine candidate MVA85A showed that although this vaccine elicits significant numbers of IFNg-secreting CD4 T cells, it does not lead to enhanced protection against infection.
Furthermore, while CD4 T cells are clearly important for TB control in humans, inherited mutations in components of the IFNg signaling pathway are not generally associated with susceptibility to M. tuberculosis but rather with enhanced susceptibility to non-tuberculosis mycobacteria.
Finally, transfer of IFNg-deficient T cells into mice can control Mtb infection to an extent.
locks the lambda phage into its lysogenic cycle, permitting transcription of λcI only through positive transcriptional regulation
the bacterial response to DNA damage. In uninduced cells, LexA, acts as a repressor of more than 40 genes , including recA and lexA. Upstream from each gene or operon, LexA binds to an operator sequence, called an SOS box. But in induced cells, damage to the DNA creates regions of single-stranded DNA that promotes the conversion of inactive RecA to an active form that can facilitate the otherwise latent capacity of LexA to auto-digest
for phage λ, the ‘switch’ from lysogenic growth to lytic growth is regulated by the SOS response.
Superinfection exclusion (Sie) systems are proteins that block the entry of phage DNA into host cells, thereby conferring immunity against specific phages. These proteins are predicted to be membrane anchored or associated with membrane components.
Coliphage T4 Sie systems
well-characterized virulent phage, has two Sie systems encoded by imm and sp. These systems cause rapid inhibition of DNA injection into cells, preventing subsequent infection by other T-even-like phages
Imm protein of Coliphage T4
Superinfection exclusion protein that prevents the transfer of phage DNA into the bacterial cytoplasm by changing the conformation of the injection site. Imm has two non-conventional transmembrane domains and is predicted to be localized to the membrane, but Imm alone does not confer complete phage immunity and must be associated with another membrane protein to exert its function and achieve complete exclusion
Sp protein of Coliphage T4
Superinfection exclusion protein that inhibits the activity of the T4 lysozyme (which is encoded by gp5), thereby presumably preventing the degradation of peptidoglycan and the subsequent entry of phage DNA. The T4 lysozyme is found at the extremity of the tail and creates holes in the host cell wall, facilitating the injection of phage DNA into the cell
Sp traps the DNA between the peptidoglycan layer and the outer membrane.
Restriction Modification Systems
When unmethylated phage DNA enters a cell harbouring a R–M system, it will be either recognized by the restriction enzyme and rapidly degraded or, to a lesser extent, methylated by a bacterial methylase to avoid restriction, therefore leading to the initiation of the phage’s lytic cycle. The fate of phage DNA is determined mainly by the processing rates of these two enzymes. As the restriction enzyme is often more active than the methylase, the incoming phage DNA is usually cleaved, although the host DNA is always protected by the methylase activity.
How have phages evolved to deal with R-M systems?
One strategy is the absence of endonuclease recognition sites in their genomes through the accumulation of point mutations
some phages have overcome R–M systems through the acquisition of the cognate methylase gene in their genomes
T4 phage genome has hydroxymethylcytosine (HMC) instead of the cytosine that is found in the host DNA. This modification allows phage T4 DNA to be impervious to R–M systems that recognize specific sequences containing a cytosine
Abi systems in bacteria
Abortive infection systems lead to the death of the infected cell. Typically, these Abi systems target a crucial step of phage multiplication such as replication, transcription or translation.
Toxin–antitoxin (TA) systems
a toxic molecule is produced by the cell and neutralized by the antitoxin product. The expression of these molecules is tightly controlled and varies from system to system but often involves promoter repression or the use of a specific transcriptional terminator. When the balance between the two regulatory halves is altered, the toxin is released and the bacterium dies.
the nuclease responsible for the first step in homology-directed DSB repair in gram-positive bacteria
Neutrophil enzymes released with NETs
Myeloperoxidase and Neutrophil Elastase