Abstract
The product of the meiosis-expressed gene 1 (MEIG1) is found in the cell bodies of spermatocytes and recruited to the manchette, a structure unique to elongating spermatids, by Parkin co-regulated gene (PACRG). This complex is essential for targeting cargo to the manchette during sperm flagellum assembly. Here we show that MEIG1 adopts a unique fold that provides a large surface for interacting with other proteins. We mutated 12 exposed and conserved amino acids and show that four of these mutations (W50A, K57E, F66A, Y68A) dramatically reduce binding to PACRG. These four amino acids form a contiguous hydrophobic patch on one end of the protein. Furthermore, each of these four mutations diminishes the ability of MEIG1 to stabilize PACRG when expressed in bacteria. Together these studies establish the unique structure and key interaction surface of MEIG1 and provide a framework to explore how MEIG1 recruits proteins to build the sperm tail.
Highlights
meiosis-expressed gene 1 (MEIG1)′ s function in spermiogenesis has been further studied in our laboratory
Mouse MEIG1 adopts a unique fold with a large solvent exposed surface area
The MEIG1 protein is well-behaved in isolation and remains a monomer at high concentration based on both the elution profile from size exclusion chromatography as well as the narrow linewidths observed in the 2D 15N-HSQC spectrum (Fig. 1)
Summary
MEIG1′ s function in spermiogenesis has been further studied in our laboratory. Using yeast two-hybrid screen, we found that Parkin co-regulated gene (PACRG) to be a major binding partner[8]. Based on this structure we mutated 12 solvent exposed amino acids in MEIG1 protein that may mediate interaction with other proteins. These 12 amino acids were mutated as described in supplemental Table 1, and binding strength between PACRG with wild-type and mutated MEIG1 proteins was tested by direct yeast two hybrid experiments.
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