| 1994 |
Porcine zona pellucida glycoprotein ZP4 amino acid sequence was determined by peptide mapping and cDNA cloning, revealing two peptide isoforms (128 and 133 amino acids) with two potential N-linked glycosylation sites, five conserved cysteine residues, and sequence similarity to mouse ZP2; ZP4 and ZP2 appear to be derived from a common parent polypeptide by proteolytic cleavage. |
Peptide mapping, cDNA cloning, sequence analysis |
Journal of reproduction and fertility |
Medium |
8182597
|
| 1998 |
Porcine ZPB (ortholog of mouse ZP1) and ZPC (ortholog of mouse ZP3) form hetero-oligomeric complexes that bind to boar sperm membrane vesicles with high affinity; neither free ZPB nor free ZPC alone binds sperm membranes, demonstrating that heterocomplex formation is required for sperm-zona binding activity. |
Solid phase binding assays, size exclusion chromatography, Western blot, biotin-labeling competition assays |
The Journal of biological chemistry |
High |
9516448
|
| 2003 |
Porcine ZPB and ZPC C-terminal residues were identified (Ala-462 and Ser-332, respectively) by mass spectrometry, revealing that ZPB is processed at its furin consensus site whereas ZPC is processed N-terminal to the furin consensus site; disulfide bond analysis within the ZP domains suggested the ZP domain consists of two subdomains. |
Mass spectrometry of C-terminal polypeptide fragments |
Biochemical and biophysical research communications |
Medium |
12878193
|
| 2007 |
Recombinant bovine ZP3 and ZP4 co-expressed in Sf9 insect cells form a hetero-complex that inhibits bovine sperm-ZP binding; neither ZP3 nor ZP4 alone shows inhibitory activity. The N-terminal and trefoil domains of ZP4 are dispensable for formation of the sperm-binding active ZP3/ZP4 complex, and the molecular interactions between ZP3 and ZP4 are conserved between bovine and porcine systems. |
Baculovirus-Sf9 cell expression, sperm-ZP competitive inhibition assay, immunoprecipitation, deletion mutagenesis |
The FEBS journal |
High |
17894824
|
| 2010 |
Human ZP4 expressed in transgenic mice is incorporated into the zona pellucida matrix with molecular mass similar to native isoforms, but is not sufficient to confer human sperm binding to mouse eggs, indicating that additional zona proteins are required for species-selective human gamete recognition. |
Transgenic mouse model, sperm binding assay, molecular mass analysis |
Reproduction (Cambridge, England) |
High |
21173071
|
| 2011 |
In porcine ZP, native ZP4 (nZP4) — but not recombinant ZP4 — is necessary for the sperm-binding activity of the ZP3/ZP4 complex toward porcine sperm, demonstrating that carbohydrate structures on ZP4 are responsible for porcine sperm-binding activity of the complex; ZP3 and ZP4 reciprocally co-precipitate each other via recombinant N-terminal tags. |
Competitive sperm-binding inhibition assay, co-immunoprecipitation, baculovirus-Sf9 expression |
Zygote (Cambridge, England) |
High |
22008510
|
| 2015 |
The hinge region (Arg-32 to Arg-160) of bovine ZP3 is a ZP4-binding site; N-glycosylation at Asn-146 (but not Asn-124) in this region is required for the sperm-binding activity of the ZP3/ZP4 complex, although it does not prevent ZP4 interaction per se. |
Baculovirus-Sf9 expression, ZP3 deletion and point mutants, co-expression, sperm-ZP binding inhibition assay |
Biomolecules |
High |
26610590
|
| 2019 |
ZP4 ablation in rabbits (CRISPR KO) produced a zona pellucida that was significantly thinner, more permeable, and more disorganized/fenestrated; ovulation, fertilization, and in vitro development were unaffected, but in vivo preimplantation development was severely impaired, demonstrating that ZP4 is required for structural integrity of the zona pellucida necessary for embryo protection in vivo. |
CRISPR-Cas9 knockout rabbit model, electron microscopy of ZP structure, permeability assays, in vivo/in vitro fertility assays |
eLife |
High |
31635692
|
| 2019 |
ZP4 is expressed in immature human fetal enterocytes and is regulated by Toll-like receptor 2 and 4 signaling; knockdown of ZP4 in fetal intestinal cells altered IL-8 mRNA expression in response to IL-1β, implicating ZP4 in innate immune signaling in the developing intestine. |
Transcription profiling, ZP4 knockdown in H4 fetal intestinal cell line, IL-8 mRNA measurement |
American journal of physiology. Gastrointestinal and liver physiology |
Low |
31314571
|
| 2021 |
Zp4 knockout female rats (CRISPR-Cas9) showed normal ZP morphology and were completely fertile, with no defects in oogenesis, ovulation, fertilization, or pup production, demonstrating that Zp4 is fully dispensable for female fertility in rats. |
CRISPR-Cas9 KO rat model, PAS staining, superovulation, in vitro fertilization, natural mating |
Biology of reproduction |
High |
33709118
|
| 2021 |
A mutant ZP1 protein in rats is intracellularly sequestered and interacts with wild-type ZP3 or ZP4 (shown in 293T cells), impeding incorporation of ZP3 and ZP4 into the zona pellucida, resulting in zona absence and female infertility. |
Homologous rat KI model, 293T cell co-expression, co-immunoprecipitation, immunofluorescence |
Biology of reproduction |
Medium |
33624742
|
| 2021 |
The N-terminal region (Lys-25 to Asp-136) and middle region (Ser-290 to Lys-340) of bovine ZP4 contain sperm-binding sites, as demonstrated by binding of sperm to ZP4-coated plates and systematic deletion mutagenesis; ZP4 harbors the major potential sperm-binding sites among bovine ZP glycoproteins, but multivalent complex formation is required for full activity. |
ZP4-coated plate solid-phase binding assay, deletion mutagenesis, Sf9-expressed recombinant proteins |
PloS one |
Medium |
34242308
|
| 2021 |
ZP4 mutations identified in human patients (whole-exome sequencing) reduced ZP4 secretion in vitro when mutant plasmids were transfected into cells, linking ZP4 loss-of-function to abnormal (thin and irregular) zona pellucida and female infertility. |
Whole-exome sequencing, in vitro plasmid expression/secretion assay in transfected cells |
Journal of clinical pathology |
Medium |
33461974
|