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Fig. 1 | BMC Molecular Biology

Fig. 1

From: Molecular analysis of NPAS3 functional domains and variants

Fig. 1

NPAS3 and ARNT interact through their bHLH and PAS domains. a Ideogram of NPAS3 constructs used. Exon numbering relative to NPAS3 transcript variant 1 corresponding to NPAS3 isoform 1 (933 aa). Only coding regions of exons are depicted. Domain constructs and variant numbering are relative to isoform 1. NPAS3 isoform 2 (901 aa) was also characterized. Red bHLH domain, Blue PAS domains, Green TAD (C-terminal transactivation domain). b Western blots of HaloTag pull-downs demonstrating that all tested isoforms of NPAS3 and ARNT are able to robustly interact, which is not observed with the empty HaloTag vector. Bands in the top panel of pull-down lane of ‘prey’ protein, indicating physical association of HA-NPAS3 isoforms with the HaloTag-ARNT construct. Middle and bottom panels are results for the HaloTag-clone and HaloTag-empty constructs. As HaloTag constructs are covalently linked to the resin, they will be depleted in the pull-down sample, demonstrating functionality of the HaloTag. Molecular weight of HaloTag-ARNT, 121 kDa, HaloTag construct with no cDNA: 34 kDa, Full-length NPAS3: 101 kDa. c Reciprocal HaloTag pull down data using HaloTag-NPAS3 constructs and untagged ARNT isoform 1. Molecular weight of HaloTag-NPAS3, 135 kDa. d HaloTag-NPAS3 pull-down of endogenously expressed ARNT. All pull-downs have been performed at least twice

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