VPS16B and VPS33B distribution in tissue and cells lysates. (A) VPS16B is found in various tissues. A ready-to-use PVDF membrane with 11 human tissue lysates (Imgenex IMB-120) was probed with anti-VPS16B mouse serum, and anti-GAPDH antibody as a loading control. (B) VPS16B and VPS33B are present in both megakaryocyte (MK) and platelet lysates (reduced 10% SDS-PAGE). The latter were probed with anti-VPS16B mouse serum, anti-VPS33B internal as well as N-terminal antibodies, along with actin as a loading control. VPS16B and actin were probed and developed simultaneously (top panel). VPS33B is detected in platelets using the internal but not N-terminal antibody. (C) VPS16B and VPS33B are primarily found in the cytosol rather than the membrane in fractionated Dami cell lysates when probed with mouse anti-VPS16B and rabbit anti-VPS33B serum, respectively. (D) Platelets from a patient with ARC syndrome containing a homozygous nonsense mutation in exon 8 of C14orf133 (VIPAR) are devoid of VPS16B (left panel; VPS16B null PLT) and contain significantly reduced VPS33B (right panel; VPS16B null PLT), as shown by immunoblotting.