Isolation and functional characterization of single domain antibody modulators of Caspase-3 and apoptosis.
Apoptosis, or programmed cell death, is an essential process affecting homeostasis of cell growth, development, and the elimination of damaged or dangerous cells. Inappropriate cell death caused by oxidative stress has been implicated in the development of neurodegenerative diseases such as Alzheimer's, Parkinson's, and stroke. On the other hand, a defect in the cell death process leads to the development of cancer. For example, the main player of apoptosis, p53, is defective in many of the human cancers. Apoptosis is regulated by the interplay of pro-apoptotic and anti-apoptotic proteins from the Bcl-2 family and caspases. In particular, specific modulators of the activity of Caspase 3 could be very important for the development of therapies for diseases such as neurodegeneration and cancer. In this study, two V(H)Hs specific to Caspase 3 (VhhCasp31 and VhhCasp32) were isolated from a heavy chain antibody variable domain (V(H)H) phage display library and tested for their apoptosis-modulating effects. While VhhCasp31 was found to be antagonistic towards Caspase 3, VhhCasp32 was agonistic. Furthermore, when expressed as intrabodies in SHSY-5Y neuroblastoma cells, VhhCasp31 rendered cells resistant to oxidative-stress-induced apoptosis, whereas VhhCasp32 resulted in apoptosis. These V(H)H antagonist and agonist of apoptosis could have potential for the development of therapeutics for neurodegenerative diseases and cancer, respectively.
McGonigal
Katrina
K
Department of Chemistry and Biochemistry, University of Windsor, Windsor, ON, Canada N9B 3P4.
Tanha
Jamshid
J
Palazov
Elitza
E
Li
Shenghua
S
Gueorguieva-Owens
Deyzi
D
Pandey
Siyaram
S
D000595:Amino Acid Sequence; D000906:Antibodies / Q000737:chemistry / Q000302:isolation & purification*; D017209:Apoptosis / Q000276:immunology*; D015153:Blotting, Western; D053148:Caspase 3 / Q000276:immunology* / Q000302:isolation & purification; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D006801:Humans; D007143:Immunoglobulin Heavy Chains / Q000737:chemistry / Q000302:isolation & purification*; D007135:Immunoglobulin Variable Region / Q000737:chemistry / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D018384:Oxidative Stress; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000302:isolation & purification; D014162:Transfection
D000906:Antibodies; D007143:Immunoglobulin Heavy Chains; D007135:Immunoglobulin Variable Region; D011994:Recombinant Proteins; D053148:Caspase 3