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Fröhlich Archangelo, Leticia (2006): Functional characterization of the CATS gene with respect to its role in normal hematopoiesis and in leukemia. Dissertation, LMU München: Faculty of Medicine
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Abstract

The Clathrin Assembly Lymphoid Myeloid leukemia gene (CALM) was first identified as the fusion partner of AF10 in the t(10;11)(p13;q14) translocation. The CALM/AF10 fusion protein plays a crucial role in t(10;11)(p13;q14) associated leukemogenesis. Using the N-terminal half of CALM as a bait in a yeast two-hybrid screen a novel protein named CATS (CALM interacting protein expressed in thymus and spleen) was identified as CALM interacting partner. Multiple tissue Northern blot analysis showed predominant expression of CATS in lymphoid tissues. CATS codes for two protein isoforms of 238 and 248 amino acids. The interaction between CALM and CATS was confirmed by co-immunoprecipitation and colocalization experiments. The CATS interaction domain of CALM was mapped to amino acids 221 to 294 of CALM. This domain is contained in the CALM/AF10 fusion protein. CATS localizes to the nucleus and shows a preference for nucleoli. Expression of CATS was able to markedly increase the nuclear localization of CALM and of the leukemogenic fusion protein CALM/AF10. This effect of CATS seems to be stronger on CALM/AF10 than on CALM. Several monoclonal antibodies against the C-terminus of human CATS were generated. These antibodies recognize both the human and the murine CATS protein. Western blot analyses showed that CATS is strongly expressed in different human leukemia, lymphoma and tumor cell lines but not in resting T-cells. High CATS expression in proliferating cells as well as its nucleolar localization suggest a role of CATS in the control of cell proliferation. In order to gain further insight into CATS function we used CATS as a bait in a yeast two-hybrid screen. Several CATS interacting proteins with apparently unrelated function were identified. Interestingly, on closer scrutiny these proteins could be associated with three key regulatory pathways: signaling, apoptosis and cell cycle control. We discuss in detail the biological relevance of the CATS interaction with the two apoptosis-associated proteins HAX1 and SIVA, the cell cycle regulator KIS and the CALM interacting ribonucleoprotein PCBP1. Our results indicate that the subcellular localization of CALM and CALM/AF10 could depend in part on the presence of CATS with a greater fraction of CALM or CALM/AF10 being present in the nucleus of cells with high CATS expression (e.g. lymphoid cells have high CATS expression). Moreover we provide evidences that CATS function might be tightly linked to cancer initiation and/or progression. The CALM-CATS interaction might thus play an important role in CALM/AF10 mediated leukemogenesis.