Abstract
Fertilization occurs as the culmination of multi-step complex processes. First, mammalian spermatozoa undergo the acrosome reaction to become fusion-competent. Then, the acrosome-reacted spermatozoa penetrate the zona pellucida and adhere to and finally fuse with the egg plasma membrane. IZUMO1 is the first sperm protein proven to be essential for sperm-egg fusion in mammals, as Izumo1 knockout mouse spermatozoa adhere to but fail to fuse with the oolemma. However, the IZUMO1 function in other species remains largely unknown. Here, we generated Izumo1 knockout rats by CRISPR/Cas9 and found the male rats were infertile. Unlike in mice, Izumo1 knockout rat spermatozoa failed to bind to the oolemma. Further investigation revealed that the acrosome-intact sperm binding conceals a decreased number of the acrosome-reacted sperm bound to the oolemma in Izumo1 knockout mice. Of note, we could not see any apparent defects in the binding of the acrosome-reacted sperm to the oolemma in the mice lacking recently found fusion-indispensable genes, Fimp, Sof1, Spaca6, or Tmem95. Collectively, our data suggest that IZUMO1 is required for the sperm-oolemma binding prior to fusion at least in rat.
Highlights
In sexual reproduction, the binding and fusion between the plasma membranes of spermatozoa and eggs are fundamental prerequisites for successful fertilization
We confirmed that Izumo1−7/−7 (KO) rats that Izumo1 mRNA derived from the mutant allele were transcribed in the testis (Figure 1C)
When Izumo1wt/−7 (Het) or Izumo1 KO male rats were individually caged with WT females for 2 months, all Izumo1 Het males sired pups, whereas Izumo1 KO males did not produce any pups despite the normal mating behavior indicated by formation of the copulation plugs (Figure 1E)
Summary
The binding and fusion between the plasma membranes of spermatozoa and eggs are fundamental prerequisites for successful fertilization. Several candidate sperm proteins involved in sperm-egg fusion have been identified using enzyme inhibitors and antibodies that block the in vitro fertilization (IVF) (Okabe, 2018). When their functions were challenged using the gene knockout (KO) mice, IZUMO1 for Sperm-Oolemma Adhesion only IZUMO1 was found indispensable for the sperm-egg fusion (Okabe, 2018; Fujihara et al, 2020; Lamas-Toranzo et al, 2020; Noda et al, 2020). The IZUMO1-JUNO, ligand-receptor interaction exists in mice and humans in a species-specific manner, implying that the two molecules might govern the gamete incompatibility among species (Bianchi and Wright, 2015)
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