TLR4 - toll like receptor 4 |Elisa - Clia - Antibody - Protein

Background

Toll-like receptor 4 (TLR4) is a member of the toll-like receptor family of proteins, which play crucial roles in the innate immune system. TLR4 is well-known for its capacity to detect lipopolysaccharide (LPS), a component of the outer membrane of Gram-negative bacteria, which is a pathogen-associated molecular pattern (PAMP) that signals the presence of bacterial infections. Upon binding to LPS, TLR4 triggers a cascade of immune responses, including the activation of nuclear factor-kappa B (NF-κB) and the production of pro-inflammatory cytokines. These responses lead to a rapid immune reaction, making TLR4 essential for the early detection and management of bacterial infections.

TLR4 is expressed in various cell types, including macrophages, dendritic cells, epithelial cells, and endothelial cells. The receptor is unique among TLRs due to its ability to signal through two distinct adaptor pathways: MyD88-dependent and TRIF-dependent pathways, which allows TLR4 to activate a broad array of immune responses. While critical in defense against infection, excessive or dysregulated TLR4 activation can contribute to chronic inflammation, autoimmune disorders, and septic shock, making TLR4 a target of interest for both enhancing and dampening immune responses therapeutically.

Protein Structure

TLR4 is a type I transmembrane protein with approximately 839 amino acids. It consists of three primary regions: an extracellular domain, a single transmembrane domain, and an intracellular Toll/IL-1 receptor (TIR) domain.

Extracellular Domain:

• The extracellular region of TLR4 contains multiple leucine-rich repeats (LRRs), which form a horseshoe-shaped solenoid. This domain is responsible for binding to ligands, including LPS.
• To recognize LPS, TLR4 forms a complex with an accessory protein called MD-2. MD-2 binds directly to LPS and, upon binding, brings two TLR4 molecules together to form a dimer. This dimerization is essential for initiating downstream signaling.
• The LRR region also contains several glycosylation sites, which stabilize the receptor structure and are crucial for its ligand-binding capacity.

Transmembrane Domain:

• The transmembrane domain is a single-pass region that anchors TLR4 within the cell membrane. This positioning allows the receptor to interact with extracellular or surface-bound PAMPs like LPS.
• The transmembrane domain itself does not play a role in ligand recognition but is essential for stabilizing the receptor within the membrane and maintaining the correct orientation for signaling.

Intracellular (TIR) Domain:

• The intracellular TIR domain is responsible for transmitting signals after TLR4 dimerization. Upon activation, the TIR domain recruits adaptor molecules, initiating downstream signaling.
• TLR4 uniquely activates two adaptor proteins, MyD88 and TRIF, allowing it to signal through both MyD88-dependent and TRIF-dependent pathways. This dual signaling capability distinguishes TLR4 from other TLRs and enables it to produce both immediate and delayed immune responses.
• The TIR domain is critical for activating NF-κB, IRF3, and MAP kinase pathways, which are involved in the production of inflammatory cytokines and type I interferons.

Classification and Subtypes

TLR4 belongs to the toll-like receptor family and is classified as part of the class I transmembrane proteins due to its single-pass transmembrane domain and extracellular LRRs. TLR4 does not have distinct subtypes but functions in association with various co-receptors and adaptors that modulate its activity and signaling outcomes.

Although TLR4 itself is unique, its functions are closely regulated by other molecules, including MD-2 (for LPS recognition) and CD14, which assists in the delivery of LPS to the TLR4/MD-2 complex. These accessory molecules modify TLR4 activity, enhancing its ability to respond to different microbial components and even endogenous ligands under certain conditions.

Function and Biological Significance

TLR4 has several important functions in the immune system, largely centered around its role in detecting Gram-negative bacteria and initiating inflammation:

LPS Recognition and Immune Activation:

• TLR4 is the primary sensor for LPS, a major component of Gram-negative bacterial cell walls. Upon LPS recognition, TLR4 forms a complex with MD-2 and dimerizes to initiate signaling. This process activates both MyD88-dependent and TRIF-dependent pathways, leading to the production of pro-inflammatory cytokines (like IL-6, TNF-α, and IL-1β) and type I interferons.
• This activation results in the recruitment and activation of additional immune cells, including neutrophils and macrophages, which play crucial roles in pathogen clearance.

MyD88-Dependent Pathway:

• The MyD88-dependent pathway is the primary pathway for most TLRs, leading to the rapid production of inflammatory cytokines through NF-κB and MAP kinase signaling. This pathway helps initiate an immediate immune response, essential for early defense against infection.
• MyD88 recruitment to TLR4 triggers a series of signaling events, including the activation of IRAK and TRAF6, which ultimately leads to NF-κB activation and cytokine production.

TRIF-Dependent Pathway:

• Unlike most other TLRs, TLR4 can also signal through the TRIF-dependent pathway, which produces a delayed but sustained response by inducing type I interferons (such as IFN-β) and promoting the maturation of dendritic cells.
• TRIF signaling is essential for stimulating the adaptive immune response, as it increases antigen presentation and enhances T-cell activation.
• This pathway is activated by TLR4 engagement in endosomes, where TLR4 can be internalized and signal through TRIF to activate IRF3 and produce type I interferons.

Role in Adaptive Immunity:

• TLR4 signaling promotes antigen presentation and dendritic cell maturation, bridging the innate and adaptive immune responses. Through cytokine production and the activation of co-stimulatory molecules, TLR4 influences the strength and duration of adaptive immune responses, particularly the activation of T-cells and B-cells.

Clinical Issues

Given its prominent role in immune responses, TLR4 is implicated in various clinical conditions:

Septic Shock:

• In cases of severe bacterial infection, overactivation of TLR4 by LPS can lead to septic shock, characterized by widespread inflammation, vascular permeability, and multiorgan failure. Septic shock results from the massive release of cytokines and is associated with high mortality.
• Therapeutic strategies aimed at blocking TLR4 or inhibiting its downstream signaling have been explored to reduce inflammation in sepsis.

Autoimmune Diseases:

• TLR4 has been implicated in autoimmune diseases like rheumatoid arthritis, systemic lupus erythematosus (SLE), and multiple sclerosis. Endogenous ligands, such as heat-shock proteins and components of necrotic cells, can trigger TLR4 inappropriately, leading to chronic inflammation and autoimmunity.
• In autoimmune settings, TLR4 signaling may contribute to tissue damage and exacerbate disease progression, highlighting TLR4 as a target for therapeutic intervention in autoimmune disorders.

Chronic Inflammatory Diseases:

• TLR4’s role in inflammation has implications for chronic inflammatory diseases, including atherosclerosis and inflammatory bowel disease (IBD). In atherosclerosis, TLR4 activation by modified LDL or other endogenous ligands promotes inflammation within blood vessels, contributing to plaque formation.
• In IBD, TLR4’s response to bacterial antigens can exacerbate inflammation within the gastrointestinal tract, worsening disease symptoms.

Cancer:

• TLR4’s role in cancer is complex. In some cases, TLR4 activation in tumor cells or the tumor microenvironment promotes inflammation, aiding in tumor progression. TLR4 can induce immunosuppressive cytokines, which allow cancer cells to evade immune surveillance.
• Conversely, TLR4 activation can stimulate immune responses against tumors, making it a potential target for immunotherapy. Modulating TLR4’s activity in the tumor microenvironment may help balance its pro- and anti-tumor effects.

Therapeutic Targeting:

• Given TLR4’s involvement in numerous diseases, it has become a target for drug development. TLR4 antagonists are being developed for use in sepsis, inflammatory diseases, and cancer, while TLR4 agonists are investigated as adjuvants in vaccines to enhance immune responses.

Summary

TLR4 is an essential receptor in the innate immune system, recognizing LPS from Gram-negative bacteria and initiating pro-inflammatory responses. Its structure, featuring an extracellular LRR domain, a transmembrane domain, and an intracellular TIR domain, enables it to detect microbial components and signal through both MyD88-dependent and TRIF-dependent pathways. These pathways coordinate immediate and sustained immune responses, making TLR4 crucial for early bacterial defense and immune activation.

Clinically, TLR4’s activation is implicated in various diseases, from sepsis to autoimmune and chronic inflammatory conditions. Overactivation can lead to excessive inflammation, as seen in septic shock, while aberrant TLR4 signaling is associated with autoimmune disorders and cancer. TLR4’s role as both a mediator of protective immunity and a contributor to pathological inflammation highlights its significance as a therapeutic target for modulating immune responses across diverse clinical conditions.