Figure S4.
The interaction of ZYG11B and IFT46 is not based on a Gly/N-degron-based mechanism. (A, C, and E) A cycloheximide chase (CHX) assay of TALPID3, BBS4, and SEPTIN2 was performed in ORF10-expressing and empty vector control HEK293T cells. Samples were taken at 0, 1, 2, and 4 h after the addition of CHX. The TALPID3, BBS4, SEPTIN2, and ORF10 protein levels were detected by immunoblotting using TALPID3, BBS4, SEPTIN2 antibodies, and HA antibody. GAPDH served as a loading control. (B, D, and F) Quantification of the TALPID3, BBS4, and SEPTIN2 levels in (A, C, and E). The protein level of TALPID3, BBS4 and SEPTIN2 was quantified using Odyssey software. (n = 3 independent experiments). Data are presented as the mean ± SEM. Statistical analysis was performed with two-tailed unpaired Student’s t test. (G) ZYG11B physically interacts with IFT46. MYC-IFT46 and ZYG11B-FLAG (or empty vector) were co-transfected into HEK293T cells. 24 h after transfection, cells were collected for immunoprecipitation (IP) with an anti-FLAG antibody, followed by analysis using anti-FLAG and anti-MYC antibodies. (H) ORF10 could not interacts with IFT46. MYC-IFT46 and ORF10-HA (or empty vector) were co-transfected into HEK293T cells. 24 h after transfection, cells were collected for immunoprecipitation (IP) with an anti-HA antibody, followed by analysis using anti-HA and anti-MYC antibodies. (I) The protein sequences of SNX11 and IFT46. (J) ZYG11B-Mut did not affect the interaction of IFT46 and ZYG11B. MYC-IFT46 and ZYG11B-FLAG (or ZYG11B-Mut-FLAG) were co-transfected into HEK293T cells. 24 h after transfection, cells were collected for immunoprecipitation (IP) with an anti-FLAG antibody, followed by analysis using anti-FLAG and anti-MYC antibodies. (K) IFT46 KR Mut did not affect the interaction of IFT46 and ZYG11B. ZYG11B-FLAG and MYC-IFT46 (or IFT46 KR Mut) were co-transfected into HEK293T cells. 24 h after transfection, cells were collected for immunoprecipitation (IP) with an anti-MYC antibody, followed by analysis using anti-FLAG and anti-MYC antibodies. Source data are available for this figure: SourceData FS4.

The interaction of ZYG11B and IFT46 is not based on a Gly/N-degron-based mechanism. (A, C, and E) A cycloheximide chase (CHX) assay of TALPID3, BBS4, and SEPTIN2 was performed in ORF10-expressing and empty vector control HEK293T cells. Samples were taken at 0, 1, 2, and 4 h after the addition of CHX. The TALPID3, BBS4, SEPTIN2, and ORF10 protein levels were detected by immunoblotting using TALPID3, BBS4, SEPTIN2 antibodies, and HA antibody. GAPDH served as a loading control. (B, D, and F) Quantification of the TALPID3, BBS4, and SEPTIN2 levels in (A, C, and E). The protein level of TALPID3, BBS4 and SEPTIN2 was quantified using Odyssey software. (n = 3 independent experiments). Data are presented as the mean ± SEM. Statistical analysis was performed with two-tailed unpaired Student’s t test. (G) ZYG11B physically interacts with IFT46. MYC-IFT46 and ZYG11B-FLAG (or empty vector) were co-transfected into HEK293T cells. 24 h after transfection, cells were collected for immunoprecipitation (IP) with an anti-FLAG antibody, followed by analysis using anti-FLAG and anti-MYC antibodies. (H) ORF10 could not interacts with IFT46. MYC-IFT46 and ORF10-HA (or empty vector) were co-transfected into HEK293T cells. 24 h after transfection, cells were collected for immunoprecipitation (IP) with an anti-HA antibody, followed by analysis using anti-HA and anti-MYC antibodies. (I) The protein sequences of SNX11 and IFT46. (J) ZYG11B-Mut did not affect the interaction of IFT46 and ZYG11B. MYC-IFT46 and ZYG11B-FLAG (or ZYG11B-Mut-FLAG) were co-transfected into HEK293T cells. 24 h after transfection, cells were collected for immunoprecipitation (IP) with an anti-FLAG antibody, followed by analysis using anti-FLAG and anti-MYC antibodies. (K) IFT46 KR Mut did not affect the interaction of IFT46 and ZYG11B. ZYG11B-FLAG and MYC-IFT46 (or IFT46 KR Mut) were co-transfected into HEK293T cells. 24 h after transfection, cells were collected for immunoprecipitation (IP) with an anti-MYC antibody, followed by analysis using anti-FLAG and anti-MYC antibodies. Source data are available for this figure: SourceData FS4.

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