Fig. 1: Identification of BRME1. | Nature Communications

Fig. 1: Identification of BRME1.

From: The BRCA2-MEILB2-BRME1 complex governs meiotic recombination and impairs the mitotic BRCA2-RAD51 function in cancer cells

Fig. 1

a Genes identified in the MEILB2 Y2H screening. Blue and red bars indicate the number of total and original clones, respectively. Genes in red indicate genes involved in this study. b Tissue-specific expressions of Brme1, Meilb2, and Gapdh (loading control) shown by RT-PCR. C2C12 is a myoblast cell line. E embryonic day. PD postnatal day. c Mapping of BRME1 peptides identified in the Y2H screening. MBD (a.a. 519–600) is the common region in all peptides. BRME1-N (a.a. 1–518) and -C (a.a. 519–605). d Y2H interactions. BRME1-N (a.a. 1–518), -C (a.a. 519–605), or -MBD (a.a. 519–600) were used as prey. MEILB2 and BRCA2-C (a.a. 2036–3329) were used as bait. e IP with the FLAG antibody from B16-F1 cells expressing MEILB2-MYC and FLAG-BRME1 truncations; F (a.a. 1–605), N (a.a. 1–518), C (a.a. 519–605), and MBD (a.a. 519–600). f Schematic of the MEILB2 sequence highlighting the recombinant protein constructs with amylose pulldown following co-expression of BRME1-MBD (a.a. 519–600) with MBP-MEILB2 α1 + 2, α1, and α2. g CD thermal denaturation, recorded as percent unfolded based on the helical signal at 222 nm, with melting temperatures estimated as shown. h, i SEC-MALS analysis. h α1 + 2 + BRME1 is a 50 kDa 2:2 complex (theoretical –48 kDa), whereas α1 + 2 forms 32 kDa dimers and 97 kDa octamers (theoretical –26 kDa and 102 kDa). Differential refractive index (dRI) profiles are overlaid with fitted molecular weights. i α1-BRME1-MBD is a 33 kDa 2:2 complex (theoretical –33 kDa), whereas α1 forms 17 kDa tetramers (theoretical –20 kDa) and α2 forms 10 kDa monomers (theoretical – 9 kDa). j SAXS P(r) distributions of α1 + 2 + BRME1, α1 + 2 (dimer), α1 + 2 (octamer), and α2 (monomer) showing maximum dimensions (Dmax) of 190 Å, 160 Å, 200 Å, and 110 Å, respectively. k SAXS ab initio models generated from 30 independent DAMMIF runs with P1 symmetry. Scale bars: 50 Å. l Schematic of α1 + 2 dimer assembly into either a 2:2 complex with BRME1 or a higher-molecular weight assembly through self-association. Summary of SEC-SAXS data are shown in Supplementary Fig. 1g. Source data are provided as a Source Data file.

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