Sperm head shaping is a key event in spermiogenesis and is tightly controlled via the acrosome–manchette network. Linker of nucleoskeleton and cytoskeleton (LINC) complexes consist of Sad1 and UNC84 ...domain–containing (SUN) and Klarsicht/ANC-1/Syne-1 homology (KASH) domain proteins and form conserved nuclear envelope bridges implicated in transducing mechanical forces from the manchette to sculpt sperm nuclei into a hook-like shape. However, the role of LINC complexes in sperm head shaping is still poorly understood. Here we assessed the role of SUN3, a testis-specific LINC component harboring a conserved SUN domain, in spermiogenesis. We show that CRISPR/Cas9-generated Sun3 knockout male mice are infertile, displaying drastically reduced sperm counts and a globozoospermia-like phenotype, including a missing, mislocalized, or fragmented acrosome, as well as multiple defects in sperm flagella. Further examination revealed that the sperm head abnormalities are apparent at step 9 and that the sperm nuclei fail to elongate because of the absence of manchette microtubules and perinuclear rings. These observations indicate that Sun3 deletion likely impairs the ability of the LINC complex to transduce the cytoskeletal force to the nuclear envelope, required for sperm head elongation. We also found that SUN3 interacts with SUN4 in mouse testes and that the level of SUN4 proteins is drastically reduced in Sun3-null mice. Altogether, our results indicate that SUN3 is essential for sperm head shaping and male fertility, providing molecular clues regarding the underlying pathology of the globozoospermia-like phenotype.
Lead ions (Pb
2+
), one form of the toxic heavy metal, have drawn significant attention due to their harmful effects on human health and the environment. Although many analytical techniques have been ...developed over the past few decades, the development of a sensitive, selective, and rapid method to detect Pb
2+
remains a challenge. In this work, we developed a sensitive surface-enhanced Raman scattering (SERS) biosensor for highly sensitive detection of Pb
2+
by using DNAzyme-modified Fe
3
O
4
@Au@Ag nanoparticles (Fe
3
O
4
@Au@Ag NPs). Firstly, the thiolated 5’-Cy3 DNA probe was modified on the surface of Fe
3
O
4
@Au@Ag NPs, which hybridized with the Pb
2+
-specific DNAzyme to form a SERS biosensor, and the Cy3 labels were used to detect Pb
2+
. In the presence of Pb
2+
, the DNAzyme cleaves the Cy3-labeled DNA probe, leading to the release of Cy3-labeled DNA probe from the Fe
3
O
4
@Au@Ag NPs. Therefore, the Raman intensity of the Cy3 labels decreases. The proposed biosensor exhibited excellent linearity in the range from 0.01 to 1.0 nM, with a limit of detection for Pb
2+
of 5 pM. It features superior selectivity to Pb
2+
over other interfering metal ions and good application in the determination of Pb
2+
in tap water and human serum samples. The SERS biosensor provides a novel‚ simple and sensitive method for detection of Pb
2+
and sheds new light on the design and synthesis of analogous SERS biosensors for the detection of other heavy metal ions.
Asthenozoospermia is a common cause of male infertility, but its etiology remains incompletely understood. We recruited three Pakistani infertile brothers, born to first-cousin parents, displaying ...idiopathic asthenozoospermia but no ciliary-related symptoms. Whole-exome sequencing identified a missense variant (c.G5408A, p.C1803Y) in DNAH17, a functionally uncharacterized gene, recessively cosegregating with asthenozoospermia in the family. DNAH17, specifically expressed in testes, was localized to sperm flagella, and the mutation did not alter its localization. However, spermatozoa of all three patients showed higher frequencies of microtubule doublet(s) 4-7 missing at principal piece and end piece than in controls. Mice carrying a homozygous mutation (Dnah17M/M) equivalent to that in patients recapitulated the defects in patients' sperm tails. Further examinations revealed that the doublets 4-7 were destabilized largely due to the storage of sperm in epididymis. Altogether, we first report that a homozygous DNAH17 missense variant specifically induces doublets 4-7 destabilization and consequently causes asthenozoospermia, providing a novel marker for genetic counseling and diagnosis of male infertility.
Here, we developed a surface-enhanced Raman scattering (SERS) sensor based on functionalized Au@Ag core–shell nanorods (Au@Ag NRs) and cascade DNAzyme amplifier (CSA) for sensitive and accurate ...determination of microRNA-21 (miRNA-21). The as-prepared SERS nanoprobes were composed of a thiol-modification hairpin probe (HP2)–functionalized Au@Ag NRs and hairpin DNAzyme (HP1-Dz). Compared with original gold nanorods, the silver shell caused an enhancement of plasmonic properties, resulting in a significant enhancement of Raman signals. In the presence of target miRNAs, the hairpin construction of HP1-Dz changed due to DNA/RNA hybridization; subsequently, the DNAzyme-catalyzed cleaving process changed, and the Raman signals of the SERS nanoprobes gradually “turned off” with time elapse because of the dissociation of the Raman reporter from the surface of Au@Ag NRs. Hence, based on this principle, the proposed SERS sensor exhibited good linearity in the range 0.5 fM to 10 nM for miRNA-21 detection with a detection limit (LOD) of 0.5 fM. The proposed SERS platform has potential application in quantitative and precise detection of miRNA-21 in human serum.
Graphical abstract
It has been found that the quality of oocytes from obese women has been compromised and subsequent embryos displayed arrested development. The compromised quality may be either due to the poor or ...rich metabolic conditions such as imbalance or excession of lipids during oocyte development. Generally, lipids are mainly stored in the form of lipid droplets and are an important source of energy metabolism. Similarly, lipids are also essential signaling molecules involved in various biological cascades of oocyte maturation, growth and oocyte competence acquisition. To understand the role of lipids in controlling the oocyte development, we have comprehensively and concisely reviewed the literature and described the role of lipid metabolism in oocyte quality and maturation. Moreover, we have also presented a simplified model of fatty acid metabolism along with its implication on determining the oocyte quality and cryopreservation for fertilization.
A novel surface-enhanced Raman scattering (SERS) analysis strategy has been designed combining Au@DTNB@Ag core
-
shell nanoparticles (DTNB attachment on gold nanoparticles, then encapsulated in Ag ...shell nanoparticles named as ADANPs) and duplex
-
specific nuclease signal amplification (DSNSA) platform. Firstly, ADANPs and magnetic substrate of Fe
3
O
4
nanoparticles were covalently attached to the 3′- and 5′- end of capture probe (CP) targeting miRNA-21. Upon the addition of target miRNA-21, these heteroduplexes were specifically cleaved by DSN and resulted in ADANPs that were released from the surface of Fe
3
O
4
nanoparticles (Fe
3
O
4
NPs). At the same time, miRNA-21 remained intact and can rehybridize another DNA probe to trigger the signal-amplifying reaction. Based on this principle, the developed SERS method exhibited good linearity in the range 0 to 1 nM for miRNA-21 with a limit of detection (LOD) of 0.084 fM and has an ability to differentiate even a single-base mismatched sequence on the target sequence or other miRNA sequence. The results provide a novel SERS method which can successfully been applied to the miRNA-21 detection in human serum.
Graphical abstract
a
shows the synthesis of Fe3O4 NPs and the conjugation of Au@DTNB@Ag NPs (ADANPs) for the detection of miRNA-21,
b
shows the operating principle of DSN-assisted signal amplification strategy for miRNA detection based on Fe3O4@CP@ADA NPs
Several genes have been reported to be involved in spermatogenesis but their functional importance in male fertility is yet needed to be elucidated. Therefore, in current research, we focused to ...explore the in vivo role of evolutionary conserved and testis-specifically expressed,
C4orf46
, gene in male mouse fertility and spermatogenesis. The expression profile of
C4orf46
is specific to testes and expressed in testes from 7 days of postpartum to onward. Thus, we generated the
C4orf46
knockout mice by utilizing CRISPR/Cas9 genome editing technology and examined gene function in spermatogenesis and fertility. Surprisingly,
C4orf46
knockout mice were completely fertile, displayed normal testes morphology, however, higher sperm contents were observed in knockout mice compared to wild type (WT) littermates. Subsequently, intact testis histology and architecture of seminiferous tubules were observed in
C4orf46
knockout and WT mice. Similarly, sperm morphology and swimming velocity of
C4orf46
knockout mice were comparable with the WT littermates. Furthermore, all type of germ cells ranging from spermatogonia to mature spermatozoa were observed in the testes and epididymis sections of
C4orf46
knockout mice suggesting that disruption of
C4orf46
did not impact spermatogenesis. Moreover, meiotic prophase I progression was normal, and each type of cell population was comparable between knockout and WT mice. Overall, finding from this research indicates that
C4orf46
is not an essential gene for fertility in mice. This study will help researchers to avoid the repetition and duplication of efforts, and to explore the genes that are indispensable for spermatogenesis and male fertility.
Multiple morphological abnormalities of the flagella (MMAF) is a genetically heterogeneous disorder leading to male infertility. Recent studies have revealed that DNAH17 variants are associated with ...MMAF, yet there is no functional evidence in support of their pathnogenicity. Here, we recruited two consanguineous families of Pakistani and Chinese origins, respectively, diagnosed with MMAF. Whole‐exome sequencing identified novel homozygous DNAH17 variants, which led to loss of DNAH17 proteins, in the patients. Transmission electron microscope analyses revealed completely disorganized axonemal structure as the predominant anomaly and increased frequencies of missings of microtubule doublet(s) 4–7 in sperm flagella of patients. Similar to those found in patients, Dnah17−/− mice also displayed MMAF phenotype along with completely disorganized axonemal structures. Clusters of disorganized microtubules and outer dense fibers were observed in developing spermatids, indicating impaired sperm flagellar assembly. Besides, we also noticed many elongating spermatids with a deformed nuclear shape and abnormal step 16 spermatids that failed to spermiate, which subsequently underwent apoptosis in Dnah17‐null mice. These findings present direct evidence establishing that DNAH17 is a MMAF‐related gene in humans and mice, extend the clinical interpretations of DNAH17 variants, and highlight an essential and complex role of DNAH17 in spermatogenesis.
Sustaining and maintaining the intricate process of spermatogenesis is liable upon hormones and growth factors acting through endocrine and paracrine pathways. The Sertoli cells (SCs) are the major ...somatic cells present in the seminiferous tubules and are considered to be the main regulators of spermatogenesis. As each Sertoli cell supports a specific number of germ cells, thus, the final number of Sertoli cells determines the sperm production capacity. Similarly, sex hormones are also major regulators of spermatogenesis and they can determine the proliferation of Sertoli cells. In the present review, we have critically and comprehensively discussed the role of sex hormones and some other factors that are involved in Sertoli cell proliferation, differentiation and maturation. Furthermore, we have also presented a model of Sertoli cell development based upon the recent advancement in the field of reproduction. Hence, our review article provides a general overview regarding the sex hormonal pathways governing Sertoli cell proliferation and development.
There are more than 2300 genes that are predominantly expressed in mouse testes. The role of hundreds of these genes has been studied in mouse spermatogenesis but still there are many genes whose ...function is unknown. Gene knockout (KO) strategy in mice is widely used for in vivo study of gene function. The present study was designed to explore the function of the four genes: Tex37, Ccdc73, Prss55 and Nxt2, which were evolutionarily conserved in eutherians. We found that these genes had a testis-enriched expression pattern in mice except Nxt2. We knocked out these genes by CRISPR/Cas9 individually and found that all the KO mice had normal fertility with no detectable difference in testis/body weight ratios, epididymal sperm counts, as well as testicular and epididymal histology from wild type mice. Although these genes are evolutionarily conserved in eutherians including human and mouse, they are not individually essential for spermatogenesis, testis development and male fertility in mice in laboratory conditions. Our report of these fertile KO data could avoid the repetition and duplication of efforts which will help in prioritizing efforts to focus on genes that are indispensable for male reproduction.
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