Structure and Function of Abnormal Spindle-Like, Microtubule Associated Protein (ASPM)

Abstract

Human autosomal recessive primary microcephaly (MCPH) is a hereditary disorder associated with a small brain and associated intellectual disability. Mutations in the gene for Abnormal Spindle-like, Microtubule Associated protein (ASPM) are the most common causes of MCPH. ASPM, a large (∼400kDa) protein, is important in mitosis. It anchors the mitotic spindle at the spindle poles and is essential for normal cytokinesis. However, the complete structure-function of ASPM is not well understood.The N-terminal domain of ASPM contains 2 tandem calponin homology (CH) domains and binds microtubules. However the location of the specific binding site is unclear. Two tandem CH domains typically bind actin, whereas single CH domains bind microtubules suggesting the CH domains of ASPM bind actin. The region following the CH domains contains 81 isoleucine-glutamine (IQ) motifs, known to bind light chains such as calmodulin. These motifs form a single alpha-helix to which the light chains bine, suggesting an overall elongated structure for ASPM.Using a combination of mass spectrometry and flow cell assays, we determined that the tandem CH domains of ASPM bind to microtubules, and not actin, suggesting that the CH domains may be the key binding site of ASPM for microtubules within the N-terminus. Negative stain electron microscopy of a central 21 IQ motif-containing domain revealed a long semi-flexible molecule, fully decorated with light chains. These data thus start to reveal the structure and function of ASPM.