Identification of Evolutionarily Conserved Md1 Splice Variants That Regulate Innate Immunity through Differential Induction of NF-кB

Abstract

Although in mammals the TLR4/myeloid differentiation factor (MD)2/CD14 complex is responsible for the recognition of bacterial LPS, and it is known that the RP105/MD1 complex negatively regulates TLR4 signaling, the evolutionary history of LPS recognition remains enigmatic. Thus, zebrafish has orthologs of mammalian TLR4 (Tlr4a and Tlr4b), RP105, and MD1, but MD2 and CD14 seem to be absent from all fish genomes available to date. In addition, and to make the story more intriguing, zebrafish Tlr4a and Tlr4b do not recognize LPS, whereas the zebrafish Rp105/Md1 complex unexpectedly participates in the regulation of innate immunity and viral resistance. In this work, we report the identification of two novel splice variants of Md1, which are expressed at similar levels as full-length Md1 in the main immune-related organs of zebrafish and are highly induced upon viral infection. One of these splice variants, which is also expressed by mouse macrophages, lacks three conserved cysteine residues that have been shown to form disulfide bonds that are crucial for the three-dimensional structure of the MD-2–related lipid recognition domain of Md1. Functional studies in zebrafish demonstrate that this evolutionarily conserved splice variant shows higher antiviral activity than full-length Md1, but reduced proinflammatory activity, due to an impaired ability to activate the master regulator of inflammation, NF-κB. These results uncover a previously unappreciated evolutionarily conserved Md1 splice variant with important functions in the regulation of innate immunity and the antiviral response in zebrafish, and point to the need for additional functional studies in mammals on this little explored molecule.

A Novel Lipopolysaccharide Recognition Mechanism Mediated by Internalization in Teleost Macrophages

“…In addition to surface receptor interaction, LPS can also activate pro-inflammatory responses by binding with intracellular receptors via internalization by electroporation ( 21 – 23 ). In the inflammasome, intracellular sensor protein activation regulates the down-stream nuclear factor-κB (NF-κB), which participates in the pro-inflammatory response ( 24 , 25 ). Nucleotide-binding oligomerization domain protein 1 (NOD1) has been implicated in the recognition of intracellular bacteria and LPS not only in mammals ( 26 ), but also in teleosts ( 27 , 28 ), suggesting that the intracellular LPS recognition pattern is similar in mammals and teleosts.…”

Inhibition of myeloid differentiation 1 specifically in colon with antisense oligonucleotide exacerbates dextran sodium sulfate‐induced colitis

“…For instance, RP105 deficiency has been shown to attenuate atherosclerosis while aggravates myocardial ischemia-reperfusion injury in mice via TLR4 signaling. 18,40,41 RP105/MD-1 not only curbs TLR4 signaling in HEK293 cell lines, neonatal rat cardiomyocytes, and myeloid cells but it also plays a promoting role in DCs maturation and B cells proliferation. 42,43 Further, considering the systemic effect of global gene deficiency, interpreting the results of global genetic deletion need caution.…”

“…16 Recent studies have shown that RP105/MD-1 attenuates inflammatory response through NF-κB signaling by blocking TLR4/MD-2 complex. 17,18 The negative regulatory role may be ascribed to heterodimerization of TLR4/MD-2 and RP105/MD-1. The complexes restrain association of the cytoplasmic TIR domains of TLR4, resulting in TLR4 signaling blockage.…”

“…Thus, RP105/MD‐1 complex serves as a negative regulatory role in TLR4 signaling . Recent studies have shown that RP105/MD‐1 attenuates inflammatory response through NF‐κB signaling by blocking TLR4/MD‐2 complex . The negative regulatory role may be ascribed to heterodimerization of TLR4/MD‐2 and RP105/MD‐1.…”

Dynamic protein expression of NF‑κB following rat intracerebral hemorrhage and its association with apoptosis

“…Certain environmental stimuli, including cytokines, free radicals, ultraviolet irradiation, ischemia, anoxia and bacterial or viral antigens, stimulate NF-κB, leading to the promotion of target gene transcription ( 15 – 17 ). The target is then recruited for physiological or pathological processes, including the inflammatory reaction, immunoreaction, cell apoptosis and free radical injury ( 18 – 21 ).…”