Mutational analysis of human papillomavirus type 16 E7 functions.

Area of Science:

• Molecular Biology
• Virology
• Oncogenesis

Background:

• Human papillomavirus type 16 (HPV16) is a major cause of cervical cancer.
• The HPV16 E7 oncoprotein is a key factor in viral oncogenesis.
• E7 protein shares homology with adenovirus E1A, suggesting conserved functional domains.

Purpose of the Study:

• To investigate the functional domains of the HPV16 E7 oncoprotein.
• To determine the role of specific amino acids and regions in E7's biological functions.
• To elucidate the structural requirements for E7's nuclear expression, transactivation, and cell transformation activities.

Main Methods:

• Construction of amino acid (AA) deletion and substitution mutants of the E7 gene.
• Expression of E7 mutants in monkey COS-1 and CV-1 cells.
• Assays for stable E7 protein expression, adenovirus E2 promoter transactivation, and rat 3Y1 cell focal transformation.

Main Results:

• Full-length E7 is necessary for stable expression; mutations in region 3 significantly reduced expression.
• Transactivation of the adenovirus E2 promoter was impaired by mutations, especially at His-2, Cys-24, and metal-binding motifs.
• Focal transformation of rat cells was abolished by mutations at His-2 and Cys-24, and significantly reduced by mutations at Cys-61 and Cys-94.

Conclusions:

• All three regions of the E7 oncoprotein are essential for its functions.
• Metal-binding motifs in region 3 are critical for maintaining the structural integrity and function of E7.
• Specific amino acid residues play crucial roles in E7's oncogenic activities, providing targets for therapeutic intervention.