Functional characterization of two RAB27A missense mutations found in Griscelli syndrome type 2

Abstract

Summary Human Griscelli syndrome type 2 (GS‐2) is characterized by partial albinism and a severe immunologic disorder as a result of RAB27A mutations. In melanocytes, Rab27A forms a tripartite complex with a specific effector Slac2‐a/melanophilin and myosin Va, and the complex regulates melanosome transport. Here, we report a novel homozygous missense mutation of Rab27A, i.e. K22R, in a Persian GS‐2 patient and the results of analysis of the impact of the K22R mutation and the previously reported I44T mutation on protein function. Both mutations completely abolish Slac2‐a/melanophilin binding activity but they affect the biochemical properties of Rab27A differently. The Rab27A(K22R) mutant lacks the GTP binding ability and exhibits cytosolic localization in melanocytes. By contrast, neither intrinsic GTPase activity nor melanosomal localization of Rab27A is affected by the I44T mutation, but the Rab27A(I44T) mutant is unable to recruit Slac2‐a/melanophilin. Interestingly, the two mutations differently affect binding to other Rab27A effectors, Slp2‐a, Slp4‐a/granuphilin‐a, and Munc13‐4. The Rab27A(K22R) mutant normally binds Munc13‐4, but not Slp2‐a or Slp4‐a, whereas the Rab27A(I44T) mutant shows reduced binding activity to Slp2‐a and Munc13‐4 but normally binds Slp4‐a.

The GTPase-deficient Rab27A(Q78L) Mutant Inhibits Melanosome Transport in Melanocytes through Trapping of Rab27A Effector Protein Slac2-a/Melanophilin in Their Cytosol

“…A similar cytosolic localization pattern of Rab27A(C219A/C221A) (Fig. 1A, and B, fifth row) as a result of mutations of geranylgeranylation sites has also been observed (18). Intriguingly, the Rab27A(C219A/C221A) mutant also strongly induced perinuclear melanosome aggregation (Fig.…”

A Heterozygous RAB27A Mutation Associated with Delayed Cytolytic Granule Polarization and Hemophagocytic Lymphohistiocytosis

“…Structural studies on related Rab27B/melanophilin (33) and Rab27A/Slp2-a (32) complexes confirm this switch 2 region of Rab27A as a binding site with Munc13-4. This is confirmed by biochemical analyses which demonstrate disruption of binding to WT Munc13-4 by mutants Rab27A p.A87P (switch 2 mutation) (36) and Rab27A p.I44T (located adjacent to switch 2 on switch 1) (38). Modeling of both the Rab27A p.I44T mutant (gray) and the patients’ p.A87P mutation (purple) predict disruption of binding to the Munc13-4 homologs, melanophilin (yellow) and Slp2-a (orange) (Figure 4).…”

Distinct Role of Rab27a in Granule Movement at the Plasma Membrane and in the Cytosol of NK Cells

“…Rab27a defects are also associated with impaired cytotoxicity [16], [17] and with poor docking of LG at the PM, as shown by electron microscopy in fixed cells [18]. However, the two RAB27A missense mutations (K22R and I44T) do not confer a dominant negative function on Rab27a in melanosome transport [19]. The corresponding mouse model is Rab27a ash mice [20].…”