Cerebral Infarction and Refractory Seizures in a Neonate with Suspected Zika Virus Infection.

Congenital Zika syndrome is a set of congenital anomalies associated with Zika virus infection during pregnancy. We present the unique case of a neonate born to a suspected Zika virus-positive mother that developed an embolic stroke and medication-resistant seizures. This report may impact the evolving definition of congenital Zika syndrome.

Loss of SED1/MFG-E8 results in altered luminal physiology in the epididymis.

SED1/MFG-E8, herein referred to as SED1, is a bimotif adhesive protein with ascribed functions in a range of cell-cell interactions, including sperm-egg binding. In the male reproductive tract, SED1 is secreted by the initial segment of the epididymis, where it coats sperm and subsequently facilitates binding to the egg zona pellucida. We have recently reported that SED1-null epididymides show an unexpected incidence of spermatic granulomas, reflecting breakdown of the epithelium and a consequent autoimmune response against sperm antigens. However, spermatic granulomas are most often manifest in the distal segments of the epididymis, whereas the bulk of SED1 is expressed in the proximal epididymis. In some models, the presence of granulomas in the distal epididymis is associated with an underlying defect in the maintenance of luminal fluid homeostasis. Herein, we report that SED1-null epididymal fluid is both hypo-osmotic and alkaline, relative to wildtype epididymal fluid. Furthermore, the SED1-null epididymal epithelium exhibits various hallmarks of disrupted fluid reabsorption and pH regulation, including altered morphology of clear cells, increased intracellular vesicles, and apical distribution of VATPase. Results indicate that the SED1-null epididymal pathologies are not the secondary consequences of defective testes or efferent ducts or of improper epididymal differentiation, unlike that seen in other epididymal models. The expression and distribution of various ion exchangers, channels, and enzymes that mediate fluid transport and pH regulation are examined in wildtype and SED1-null epididymides, and models to account for how SED1 functions in luminal fluid dynamics are discussed.

A novel role for SED1 (MFG-E8) in maintaining the integrity of the epididymal epithelium.

The epididymis is a highly convoluted tubule that connects the testis with the vas deferens, and in which mammalian sperm acquire the ability to fertilize eggs. The most proximal portion of the epididymis, or initial segment, secretes numerous factors that are critical for sperm maturation and storage. One such factor is SED1 (also known as MFG-E8) a bi-motif protein composed of two N-terminal EGF domains, the second of which contains an RGD motif, and two C-terminal discoidin domains (also known as F5/8 type C domains). Previous studies have reported that SED1 is secreted into the epididymal lumen, where it coats sperm and later facilitates sperm-egg binding. Herein, we report that SED1-null males also harbor unexpected epididymal pathologies, including detached epithelia and spermatic granulomas. We therefore examined whether SED1 has a tissue-intrinsic role in the epididymis, in addition to its role in sperm-egg adhesion. Improved fixation protocols revealed that SED1 is found in the basolateral domains of epididymal epithelial cells in vivo, and similarly, SED1 is secreted both apically and basally from polarized epididymal cells in vitro. The basolateral distribution of SED1 suggests that it may play a novel role in epididymal cell adhesion. Consistent with this, in vitro assays showed that SED1 supports epididymal cell adhesion via RGD binding to alphaV integrin receptors on epididymal epithelial cells. Finally, epididymal cells from SED1-null males showed reduced adhesion in vitro, a phenotype that can be rescued with exogenous SED1. These results suggest that SED1 facilitates epididymal cell adhesion, and that its loss leads to breakdown of the epididymal epithelium and consequent development of spermatic granulomas.

SED1/MFG-E8: a bi-motif protein that orchestrates diverse cellular interactions.

MFG-E8 was initially identified as a principle component of the Milk Fat Globule, a membrane-encased collection of proteins and triglycerides that bud from the apical surface of mammary epithelia during lactation. It has since been independently identified in many species and by many investigators and given a variety of names, including p47, lactadherin, rAGS, PAS6/7, and BA-46. The acronym SED1 was proposed to bring cohesion to this nomenclature based upon it being a Secreted protein that contains two distinct functional domains: an N-terminal domain with two EGF-repeats, the second of which has an integrin-binding RGD motif, and a C-terminal domain with two Discoidin/F5/8C domains that bind to anionic phospholipids and/or extracellular matrices. SED1/MFG-E8 is now known to participate in a wide variety of cellular interactions, including phagocytosis of apoptotic lymphocytes and other apoptotic cells, adhesion between sperm and the egg coat, repair of intestinal mucosa, mammary gland branching morphogenesis, angiogenesis, among others. This article will explore the various roles proposed for SED1/MFG-E8, as well as its provocative therapeutic potential.

Milk fat globule-EGF factor 8/lactadherin plays a crucial role in maintenance and repair of murine intestinal epithelium.

Milk fat globule-EGF factor 8 (MFG-E8)/lactadherin participates in several cell surface-mediated regulatory events. Although its mRNA is present in the gut, the physiological roles of MFG-E8 in the intestinal mucosa have not been explored. Here we show that MFG-E8 was expressed in intestinal lamina propria macrophages from mice. Using a wound-healing assay, MFG-E8 was shown to promote the migration of intestinal epithelial cells through a PKCepsilon-dependent mechanism. MFG-E8 bound to phosphatidylserine and triggered reorientation of the actin cytoskeleton in intestinal epithelial cells at the wound edge. Depleting MFG-E8 in mice by administration of anti-MFG-E8 antibody or targeted deletion of the MFG-E8 gene resulted in a slowing of enterocyte migration along the crypt-villus axis and focal mucosal injury. Moreover, in septic mice, intestinal MFG-E8 expression was downregulated, which correlated with intestinal injury, interrupted enterocyte migration, and impaired restitution. Treatment with recombinant MFG-E8 restored enterocyte migration, whereas deletion of MFG-E8 impeded mucosal healing in mice with sepsis. These results suggest that a decrease in intestinal MFG-E8 impairs intestinal mucosal repair in sepsis. Together, our data indicate that MFG-E8 plays an important role in the maintenance of intestinal epithelial homeostasis and the promotion of mucosal healing and suggest that recombinant MFG-E8 may be beneficial for the treatment of bowel injuries.

Novel gamete receptors that facilitate sperm adhesion to the egg coat.

Mammalian fertilization is initiated by species-specific binding of the sperm to the zona pellucida, or egg coat. Previous studies suggested that sperm adhesion to the egg coat is facilitated, at least in part, through the binding of sperm surface beta1 ,4-galactosyltransferase I (GaIT) to glycoside chains on the egg coat glycoprotein, ZP3. Binding of multiple ZP3 oligosaccharides induces aggregation of GaIT within the sperm membrane, triggering, directly or indirectly, a pertussis toxin sensitive G-protein cascade leading to induction of the acrosome reaction. Consistent with this, spermatozoa bearing targeted deletions in GaIT are unable to bind ZP3 or undergo ZP3-dependent acrosomal exocytosis; however, unexpectedly, GaIT-null sperm are still able to bind to the egg coat. This indicates that sperm-egg binding requires at least two independent binding mechanisms; a GaIT-ZP3-independent event that mediates initial adhesion, followed by a GaIT-ZP3 interaction that facilitates acrosomal exocytosis. Our recent efforts have focused on the identification and characterization of these novel gamete receptors. One recently identified sperm protein that is required for sperm adhesion to the egg coat is SED1. SED1 is a bimotif protein composed of two Notch-like EGF repeats and two discoidin/complement F5/8 domains. SED1 is secreted by the epididymal epithelium and coats spermatozoa as they progress through the epididymis. Spermatozoa null for SED1 fail to bind the egg coat, illustrating its requirement for gamete adhesion. Interestingly, SED1 is also expressed by a variety of other epithelial tissues, where it appears to be required for epithelial morphogenesis and/or maintenance. A second novel gamete receptor has recently been identified on the coat of ovulated oocytes. This ZP3-independent, egg coat component is a high molecular weight, wheat germ agglutinin (WGA)-reactive glycoprotein that is derived from oviduct secretions and appears to participate in initial sperm adhesion. The amino acid sequence of this oviduct-derived ligand is currently being determined for the generation of peptide-specific antibodies and for the creation of knock out mice. The identification of novel gamete receptors that are required for sperm-egg binding opens up new avenues for the development of specific contraceptive strategies.

Identification of novel gamete receptors that mediate sperm adhesion to the egg coat.

Mammalian fertilization is initiated by the species-specific binding of sperm to the zona pellucida, or egg coat. Earlier studies suggested that sperm-egg adhesion in mouse is mediated by the binding of beta1,4-galactosyltransferase-I (GalT) on the sperm surface to specific glycoside ligands on the egg coat glycoprotein, ZP3. Binding of multiple ZP3 oligosaccharides induces GalT aggregation, triggering a pertussis toxin-sensitive G-protein cascade leading to induction of the acrosome reaction. Consistent with this, sperm bearing targeted deletions in GalT are unable to bind ZP3 nor undergo ZP3-dependent acrosomal exocytosis; however, GalT-null sperm are still able to bind to the egg coat. This indicates that sperm-egg binding requires at least two independent binding mechanisms: a GalT-ZP3-independent event that mediates initial adhesion, followed by a GalT-ZP3 interaction that facilitates acrosomal exocytosis. During the past few years, novel GalT-ZP3-independent gamete receptors have been identified that appear to participate in initial gamete adhesion. On such receptor is SED1, an EGF repeat and discoidin domain protein that coats sperm as they traverse through the epididymis, and which is required for sperm to bind the egg coat. Similarly, a novel egg coat ligand is present on ovulated oocytes, but not on ovarian eggs, and which also appears to function in initial sperm binding. The identification of novel gamete receptors that are required for sperm-egg binding opens up new avenues for the development of specific contraceptive strategies.

Functional genomics reveals relationships between the retrovirus-like Ty1 element and its host Saccharomyces cerevisiae.

Retroviruses and their relatives, the long terminal repeat (LTR) retrotransposons, carry out complex life cycles within the cells of their hosts. We have exploited a collection of gene deletion mutants developed by the Saccharomyces Genome Deletion Project to perform a functional genomics screen for host factors that influence the retrovirus-like Ty1 element in yeast. A total of 101 genes that presumably influence many different aspects of the Ty1 retrotransposition cycle were identified from our analysis of 4483 homozygous diploid deletion strains. Of the 101 identified mutants, 46 had significantly altered levels of Ty1 cDNA, whereas the remaining 55 mutants had normal levels of Ty1 cDNA. Thus, approximately half of the mutants apparently affected the early stages of retrotransposition leading up to the assembly of virus-like particles and cDNA replication, whereas the remaining half affected steps that occur after cDNA replication. Although most of the mutants retained the ability to target Ty1 integration to tRNA genes, 2 mutants had reduced levels of tRNA gene targeting. Over 25% of the gene products identified in this study were conserved in other organisms, suggesting that this collection of host factors can serve as a starting point for identifying host factors that influence LTR retroelements and retroviruses in other organisms. Overall, our data indicate that Ty1 requires a large number of cellular host factors to complete its retrotransposition cycle efficiently.