Increased Expression of an HIV CJ505.664 SOSIP Envelope in Plants by Modulation of ER Chaperones

The recombinant HIV SOSIP EnvelopeEnvtrimer is the leading HIV vaccine candidate but current production levels are low in mammalian cell lineslikely due to current trimer designs which include numerous disulfide bondsproteolytic cleavage and extensive glycosylationknown to induce endoplasmic reticulumERstress and cellular toxicityThis proposal will utilize the Nicotiana benthamiana pN bpplant expression system to produce the CHTFR SOSIPvEnvCHSOSIPand address whether manipulation of ER haperones will increase the host cell capacity for correct foldingdisulfide bond formation and post translational modificationThusthe unfolded protein responseUPRwill be induced by co expressing the master regulators bZIPs and bZIPtactive forms of transcription factorsand selected ER chaperonesBiPPDIPPICRT and CNXindividually and in combinationThe N bpplant platform offers an advantage for rapidly assessing cellular toxicity by phenotypic screening ofleaf browningafterdaysIn additionthe proposed studies will examine whether fusions between the anti CLEC A antibody and CHSOSIP Env molecules will enhance stability and expression levels as well as provide for a generic protein A based downstream processing strategyFinallythe suitability of a stress resistant plant hostNicotiana tabacumfor producing VHSOSIP Env at a commercial scale will be evaluated