Autophagy and ubiquitin–proteasome system contribute to sperm mitophagy after mammalian fertilization

Abstract

Maternal inheritance of mitochondria and mtDNA is a universal principle in human and animal development, guided by selective ubiquitin-dependent degradation of the sperm-borne mitochon-dria after fertilization. However, it is not clear how the 26S pro-teasome, the ubiquitin-dependent protease that is only capable of degrading one protein molecule at a time, can dispose of a whole sperm mitochondrial sheath. We hypothesized that the canonical ubiquitin-like autophagy receptors [sequestosome 1 (SQSTM1), microtubule-associated protein 1 light chain 3 (LC3), gamma-aminobutyric acid receptor-associated protein (GABARAP)] and the nontraditional mitophagy pathways involving ubiquitin-proteasome system and the ubiquitin-binding protein dislocase, valosin-containing protein (VCP), may act in concert during mam-malian sperm mitophagy. We found that the SQSTM1, but not GABARAP or LC3, associated with sperm mitochondria after fertilization in pig and rhesus monkey zygotes. Three sperm mitochondrial proteins copurified with the recombinant, ubiquitin-associated domain of SQSTM1. The accumulation of GABARAP-containing protein aggregates was observed in the vicinity of sperm mitochondrial sheaths in the zygotes and increased in the embryos treated with proteasomal inhibitor MG132, in which intact sperm mitochondri-al sheaths were observed. Pharmacological inhibition of VCP significantly delayed the process of sperm mitophagy and completely prevented it when combined with microinjection of autophagy-targeting antibodies specific to SQSTM1 and/or GABARAP. Sperm mitophagy in higher mammals thus relies on a combined action of SQSTM1-dependent autophagy and VCP-mediated dislocation and presentation of ubiquitinated sperm mitochondrial proteins to the 26S proteasome, explaining how the whole sperm mitochondria are degraded inside the fertilized mammalian oocytes by a protein recycling system involved in degradation of single protein molecules. mitochondria | mtDNA | ubiquitin | autophagy | mitophagy T he inheritance pattern of the mitochondrial genome does not follow Mendelian rules as mtDNA is predominantly or exclusively inherited from the mother in almost all eukaryotic species studied, which is referred to as maternal inheritance of mtDNA (1, 2). The proteolytic ubiquitin-proteasome system (UPS) for substrate-specific, regulated protein recycling has been implicated in the targeted degradation of paternal mi-tochondria (sperm mitophagy) in mammals and other taxa. Mammalian sperm mitochondria are already modified with ubiquitin during spermatogenesis and ultimately processed by proteasome-mediated proteolysis (3). Specific cell-permeant inhibitors of proteasomal chymotrypsin-like activity, MG132 and lactacystin, blocked the progression of sperm mitophagy after porcine fertilization, and the resumption of sperm mi-tochondrion degradation was observed once the MG132 (a reversible inhibitor) was removed from the zygotes (4). Our early study reported that sperm tails in zygotes were surrounded by lysosome-like structures, which sugge...

Mitophagy in neurodegeneration and aging

“…MUL1 can also cleave sperm-derived mitochondria in a PINK1-dependent manner in zygotes [41](detailed below). Additionally, Nix/BNIP3L can induce mitophagy in a PINK1/Parkin-independent manner [42] (also shown below). Some proteins involved in Parkin-independent mitophagy pathways, such as Ambra1, Nix, and FUNDC1, also have roles in traditional Parkin-dependent mitophagy [43–45].…”

Assisted reproductive technologies to prevent human mitochondrial disease transmission

“…In a conserved process involving concerted autophagy (mitophagy) and proteasomal degradation pathways 7 , sperm-derived mitochondria are eliminated after fertilization before the two-cell stage [8][9][10] although alternative mechanisms, including elimination of sperm mtDNA before fertilization, may operate in the mouse 11 .…”

“…In mammalian fertilization, mitophagy naturally removes sperm-borne mitochondria via a pathway that ubiquitylates mitochondrial components for proteasomal destruction 10 . Mitophagy may be restricted to cells of the male germline, as mitochondria from mouse spermatids 1 0 6 6 VOLUME 35 NUMBER 11 NOVEMBER 2017 nature biotechnology r e v i e w but not hepatocytes are degraded 9 , which is why the process would have to be modified so that carried-over mitochondria were ubiquitylated in MRT protocols.…”

Persistence and Transcription of Paternal mtDNA Dependent on the Delivery Strategy Rather than Mitochondria Source in Fish Embryos

“…Yu et al showed that the mitochondrial DNA polymerase promotes abrupt elimination of mitochondrial genomes during spermatogenesis in Drosophila [25]. Upon fertilization, proteasomal and lysosomal pathways are activated in oocytes to eliminate paternal MT and mtDNA [6,26,27]. In C. elegans, two groups have shown that paternal MTs are degraded by fertilization-induced autophagy [26,28].…”

“…In mammals, sperm-derived paternal mitochondria themselves are thought to undergo selective autophagy, which includes mitophagy. Ubiquitination of the sperm mitochondria during spermatogenesis and after-fertilization has been implicated in the targeted degradation of paternal mitochondria [27,29,31,32]. However, it is not known whether or not mtDNA degradation could be delayed in mammalian embryos because of the method of delivery of paternal mitochondrial DNA by injection .…”