Q52 — Cryopyrin Inflammasome — Western Blot of WT and Mutant Variants

Theoretical 2 Real exam question — full text reproduced under IBO's CC BY-NC-SA 4.0 license

Cryopyrin is a protein involved in the formation of enzymatically active caspase-1 from procaspase-1. p35 is a protein fragment resulting from the cleavage of procaspase-1. The conversion of pro-interleukin-1β to interleukin-1β (IL-1β) is catalyzed by caspase-1. Researchers were interested in the functional properties of wild-type cryopyrin and its two mutants, R260W and D303N. They developed two cell lines: 293-ASC-caspase-1 with a stable expression of both procaspase-1 and ASC (a protein known to interact with cryopyrin) and 293-caspase-1, with a stable expression of procaspase-1 but not ASC. They transfected cells from each cell line with one of the five vectors and then performed a Western Blot of the proteins they were interested in (Figure).

Figure. Western blot results for each cell line: (1) empty vector, (2) vector with WT cryopyrin, (3) vector with the cryopyrin R260W mutant, (4) vector with the cryopyrin D303N mutant, (5) vector with IPAF-ΔLRR, which causes autocleavage of procaspase-1 independent from all other factors. Rows show Procasp-1/p35, Pro-IL-1β/IL-1β, Cryopyrin, and ASC. Figure. Results of Western Blot for each cell line, 293-caspase-1 (left) and 293-ASC-caspase-1 (right).

Using the information and figures, determine whether the following statements are true or false.

A. Cryopyrin-dependent IL-1β production is not dependent on ASC.

B. The R260W mutant is partially active, as compared to wild type cryopyrin.

C. The D303N mutant does not contain a truncating mutation early in the gene sequence.

D. IPAF-ΔLRR served as a positive control in this experiment for each cell line.

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Question reproduced from IBO 2022, Theoretical Paper 2, licensed under CC BY-NC-SA 4.0 — attributed to the International Biology Olympiad. Open the full exam PDF · Community solutions (unofficial)