Smap1 deficiency perturbs receptor trafficking and predisposes mice to myelodysplasia
Shunsuke Kon, … , Takuro Nakamura, Masanobu Satake
Shunsuke Kon, … , Takuro Nakamura, Masanobu Satake
Published February 22, 2013
Citation Information: J Clin Invest. 2013;123(3):1123-1137. https://doi.org/10.1172/JCI63711.
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Research Article Hematology

Smap1 deficiency perturbs receptor trafficking and predisposes mice to myelodysplasia

Abstract

The formation of clathrin-coated vesicles is essential for intracellular membrane trafficking between subcellular compartments and is triggered by the ARF family of small GTPases. We previously identified SMAP1 as an ARF6 GTPase-activating protein that functions in clathrin-dependent endocytosis. Because abnormalities in clathrin-dependent trafficking are often associated with oncogenesis, we targeted Smap1 in mice to examine its physiological and pathological significance. Smap1-deficent mice exhibited healthy growth, but their erythroblasts showed enhanced transferrin endocytosis. In mast cells cultured in SCF, Smap1 deficiency did not affect the internalization of c-KIT but impaired the sorting of internalized c-KIT from multivesicular bodies to lysosomes, resulting in intracellular accumulation of undegraded c-KIT that was accompanied by enhanced activation of ERK and increased cell growth. Interestingly, approximately 50% of aged Smap1-deficient mice developed anemia associated with morphologically dysplastic cells of erythroid-myeloid lineage, which are hematological abnormalities similar to myelodysplastic syndrome (MDS) in humans. Furthermore, some Smap1-deficient mice developed acute myeloid leukemia (AML) of various subtypes. Collectively, to our knowledge these results provide the first evidence in a mouse model that the deregulation of clathrin-dependent membrane trafficking may be involved in the development of MDS and subsequent AML.

Authors

Shunsuke Kon, Naoko Minegishi, Kenji Tanabe, Toshio Watanabe, Tomo Funaki, Won Fen Wong, Daisuke Sakamoto, Yudai Higuchi, Hiroshi Kiyonari, Katsutoshi Asano, Yoichiro Iwakura, Manabu Fukumoto, Motomi Osato, Masashi Sanada, Seishi Ogawa, Takuro Nakamura, Masanobu Satake

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Figure 2

ARF6 activation and transferrin endocytosis in bone marrow cells.

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ARF6 activation and transferrin endocytosis in bone marrow cells. (A) Pr...
(A) Protein lysates were prepared from Smap1+/+ and Smap1–/– bone marrow cells and incubated with GST or GST-GGA1 coupled to glutathione-Sepharose. The bound fraction was processed for immunoblot detection by anti-ARF6–specific and anti-panARF antibodies, as indicated. An asterisk represents nonspecific bands. The amounts of ARF6 in each lysate prior to incubation with GST or GST-GGA1 were also evaluated by immunoblotting (see “Total ARF6”). (B and C) Bone marrow cells were prepared from Smap1+/+ and Smap1–/– mice and labeled with fluorescein-transferrin at 4°C. Excessive transferrin in the medium was washed away (initially bound transferrin at this time is shown as “4°C” as indicated in the top left of B), and, after incubation of cells at 37°C for the indicated time, surface-remaining transferrin was stripped off. Cells were labeled with PE-anti-Ter119 and processed for flow cytometry. The Ter119+ fraction was gated, and the transferrin-derived fluorescence intensities are displayed. Relative amounts of internalized fluorescein were measured by comparing fluorescence intensities at 0 minutes and each given time. Cells were prepared from 3 independent pairs of Smap1+/+ and Smap1–/– mice and processed for assays. Averages ± SD of internalized transferrin were calculated for each incubation time at 37°C (n = 3). *P < 0.05.