Difference between revisions of "Part:BBa K2632003"

Revision as of 04:45, 16 September 2018

A377D mutant of full length Gasdermin D

Pyroptosis is a form of lytic programmed cell death with inflammation. Recent studies reported that N-terminal of Gasdermin D acts as a effector of pyroptosis. Full length Gasdermin D is cleaved by Caspase 1 then release the PFD(pore-forming domain) which can oligomerize on the cell membrane. Formation of pore causes cell swelling, rupture of the membrane and massive leakage of cytosolic contents. We respectively fused EGFP with GSDMD-N275, GSDMD FL(full length) and L290D,Y373D,A377D mutation of GSDMD FL. Then transfect these plasmids into HEK293T cell. Microscopy of GSDMD-N275 undergoing pyroptosis, but GSDMD full length did not induce pyroptosis(Fig 1). We also test the cell viability though an ATP assay (CellTiter-Glo® Luminescent Cell Viability Assay) and demonstrate that GSDMD-N275 and mutation of GSDMD FL have different ability to induce pyroptosis(Fig 2).

<img src="" width="700px"/> <img src="" width="700px"/> </br> Figure 1. pCS2-EGFP-GSDMD FL（left）, pCS2-EGFP-GSDMD-N275（right） were transfected respectively into 293T cells. <img src="" width="700px"/> </br> Figure 2. pCS2-EGFP-GSDMD FL, pCS2-EGFP-GSDMD-N275, pCS2-EGFP-GSDMD L290D, pCS2-EGFP-GSDMD Y373, pCS2-EGFP-GSDMD A377D were transfected respectively into 293T cells. ATP-based cell viability was measured. Sequence and Features