Characterizing the Molecular Players Involved in Lipopolysaccharide Inflammatory Responses — ASN Events

Characterizing the Molecular Players Involved in Lipopolysaccharide Inflammatory Responses (#336)

Elizabeth J Thatcher 1 , Douglas T Golenbock 1
  1. U Mass Medical School, Worcester, MA, United States

The human body relies on its ability to sense invading microorganisms in order to mount an effective immune response. A primary player in the immune systems response to gram-negative bacterial invasion is lipopolysaccharide (LPS). LPS is a critical component of the outer membrane of gram-negative bacteria, and its endotoxin properties can elicit a large inflammatory response that can induce sepsis and subsequent death. Presently, we know that the binding of LPS to CD14 initiates signal transduction; however, the molecular processes immediately following this event remains unclear.

Previously, Chinese hamster ovary (CHO)-K1 cells that stably express CD14 were mutagenized and exposed to LPS. Two complementation groups were isolated from this screen, both of which fail to response to LPS stimulation but still respond to TNF stimulation. Complementation Group A was further characterized and found to contain mutations in MD-2. Soluble MD-2 has since been shown as a critical regulator of cellular responses to LPS through TLR4 signaling. Here, I have utilized whole transcriptome sequencing to characterize all mutations in Complementation Group B. I am currently using wild-type expression constructs isolated from the other complementation group to verify the gene required to induce a LPS non-responsive phenotype in Complementation Group B. After in vitro confirmation, these mutations will be incorporated into engineered mouse models in order to characterize the in vivo response of this mutation.