scholarly journals Loss of the cleaved-protamine 2 domain leads to incomplete histone-to-protamine exchange and infertility in mice

2021 ◽  
Author(s):  
Lena Arévalo ◽  
Gina Esther Merges ◽  
Simon Schneider ◽  
Franka Enow Oben ◽  
Isabelle Neumann ◽  
...  
Keyword(s):  
Dna Damage ◽  
Gene Editing ◽  
Chromatin Condensation ◽  
Mammalian Species ◽  
Complete Removal ◽  
Sperm Chromatin ◽  
Male Mice ◽  
Specific Proteins ◽  
Species Specific

Protamines are unique sperm-specific proteins that package and protect paternal chromatin until fertilization. A subset of mammalian species expresses two protamines (PRM1 and PRM2), while in others PRM1 is sufficient for sperm chromatin packaging. Alterations of the species-specific ratio between PRM1 and PRM2 are associated with infertility. Unlike PRM1, PRM2 is generated as a precursor protein consisting of a highly conserved N-terminal domain, termed cleaved PRM2 (cP2), which is consecutively trimmed off during chromatin condensation. The carboxyterminal part, called mature PRM2 (mP2), interacts with DNA and mediates chromatin hyper-condensation, together with PRM1. The removal of the cP2 domain is believed to be imperative for proper chromatin condensation and the prevention of DNA damage. Yet, the role of cP2 is not yet understood. Using CRISPR-Cas9 mediated gene editing, we generated mice lacking the cP2 domain while the mP2 is still expressed. We show that deletion of one allele of the cP2 domain is sufficient to render male mice infertile. cP2 deficient sperm show incomplete PRM2 incorporation, retention of transition proteins and a severely altered protamine ratio. During epididymal transit, cP2 deficient sperm seem to undergo ROS mediated degradation leading to complete DNA fragmentation, inviability and immotility of mature sperm. The cP2 domain therefore seems to be necessary for the complex crosstalk leading to the successive and complete removal of transition proteins and complete protamination of sperm chromatin. Overall, we present the first step towards understanding the role of the cP2 domain in paternal chromatin packaging and open up avenues for further research.

scholarly journals Role of inhibitory CDC2 phosphorylation in radiation-induced G2 arrest in human cells.

1996 ◽  
Vol 134 (4) ◽  
pp. 963-970 ◽  
Author(s):  
P Jin ◽  
Y Gu ◽  
D O Morgan
Keyword(s):  
Dna Damage ◽  
Chromatin Condensation ◽  
S Phase ◽  
Human Cells ◽  
G2 Arrest ◽  
Hela Cell Lines ◽  
Radiation Induced ◽  
G2 Delay ◽  
In Cells

The activity of the mitosis-promoting kinase CDC2-cyclin B is normally suppressed in S phase and G2 by inhibitory phosphorylation at Thr14 and Tyr15. This work explores the possibility that these phosphorylations are responsible for the G2 arrest that occurs in human cells after DNA damage. HeLa cell lines were established in which CDC2AF, a mutant that cannot be phosphorylated at Thr14 and Tyr15, was expressed from a tetracycline-repressible promoter. Expression of CDC2AF did not induce mitotic events in cells arrested at the beginning of S phase with DNA synthesis inhibitors, but induced low levels of premature chromatin condensation in cells progressing through S phase and G2. Expression of CDC2AF greatly reduced the G2 delay that resulted when cells were X-irradiated in S phase. However, a significant G2 delay was still observed and was accompanied by high CDC2-associated kinase activity. Expression of wild-type CDC2, or the related kinase CDK2AF, had no effect on the radiation-induced delay. Thus, inhibitory phosphorylation of CDC2, as well as additional undefined mechanisms, delay mitosis after DNA damage.

scholarly journals Role of HP1β during spermatogenesis and DNA replication

2019 ◽  
Author(s):  
Vijaya Charaka ◽  
Anjana Tiwari ◽  
Raj K Pandita ◽  
Clayton R Hunt ◽  
Tej K. Pandita
Keyword(s):  
Dna Damage ◽  
Dna Replication ◽  
Germ Cells ◽  
Chromatin Condensation ◽  
Genomic Stability ◽  
Replication Forks ◽  
Chromatin Protein ◽  
Male Germ Cells ◽  
Chromatin Compaction

AbstractMaintaining genomic stability in a continually dividing cell population requires accurate DNA repair, especially in male germ cells. Repair and replication protein access to DNA, however, is complicated by chromatin compaction. The HP1β chromatin protein, encoded by Cbx1, is associated with chromatin condensation but its role in meiosis is not clear. To investigate the role of Cbx1 in male germ cells, we generated testis specific Cbx1 deficient transgenic mice by crossing Cbx1flox/flox (Cbx1f/f) mice with Stra8 Cre+/− mice. Loss of Cbx1 in testes adversely affected sperm maturation and Cbx1 deletion increased seminiferous tubule degeneration and basal level DNA damage., We observed that Cbx1−/− MEF cells displayed reduced resolution of stalled DNA replication forks as well as decreased fork restart, indicating defective DNA synthesis. Taken together, these results suggest that loss of Cbx1 in growing cells leads to DNA replication defects and associated DNA damage that impact cell survival.

scholarly journals Adult hippocampal neurogenesis of mammals: evolution and life history

2008 ◽  
Vol 5 (1) ◽  
pp. 141-144 ◽  
Author(s):  
Irmgard Amrein ◽  
Hans-Peter Lipp
Keyword(s):  
Hippocampal Formation ◽  
Mammalian Species ◽  
Critical Role ◽  
Hippocampal Neurogenesis ◽  
Adult Hippocampal Neurogenesis ◽  
Mammalian Brain ◽  
Spatial Learning And Memory ◽  
Species Specific ◽  
Species Being

Substantial production of new neurons in the adult mammalian brain is restricted to the olfactory system and the hippocampal formation. Its physiological and behavioural role is still debated. By comparing adult hippocampal neurogenesis (AHN) across many mammalian species, one might recognize a common function. AHN is most prominent in rodents, but shows considerable variability across species, being lowest or missing in primates and bats. The latter finding argues against a critical role of AHN in spatial learning and memory. The common functional denominator across all species investigated thus far is a strong decline of AHN from infancy to midlife. As predicted by Altman and colleagues in 1973, this implies a role in transforming juvenile unpredictable to predictable behaviour, typically characterizing mammalian behaviour once reproductive competence has been attained. However, as only a fraction of mammalian species has been investigated, further comparative studies are necessary in order to recognize whether AHN has a common unique function, or whether it mediates species-specific hippocampal functions.

scholarly journals 215.Analysis of DNA damage induced by pro-oxidant treatment of mammalian spermatozoa in vitro

2004 ◽  
Vol 16 (9) ◽  
pp. 215
Author(s):  
L. E. Bennetts ◽  
R. J. Aitken
Keyword(s):  
Dna Damage ◽  
Oxidative Damage ◽  
Dna Sequences ◽  
Nuclear Dna ◽  
Quantitative Polymerase Chain Reaction ◽  
Mammalian Species ◽  
Dose Range ◽  
Tammar Wallaby ◽  
Sperm Chromatin ◽  
Sperm Dna

Defects in the male genome produced as a consequence of oxidative insult have been associated with decreased fertility levels, an elevated incidence of childhood cancer and dominant genetic disease in the offspring (1). The objective of this study was to determine the relative susceptibility of sperm DNA of different mammalian species to oxidative injury. We applied a highly sensitive quantitative PCR assay (QPCR) to measure gene-specific DNA damage in nuclear and mitochondrial compartments of spermatozoa treated with H2O2. Human, murine and tammar wallaby (Macropus eugenii) spermatozoa were treated with H2O2 (0–5�mM) over a 1�h period. After DNA purification, DNA damage was assessed in a nuclear and a mitochondrial fragment of DNA by quantitative polymerase chain reaction assay (QPCR). DNA damage was detected as a decrease amplification of the target sequences. In murine and human spermatozoa, mitochondrial DNA exhibited greater sensitivity to oxidative damage than nuclear DNA. Doses ranging from 0.25–5�mM H2O2 induced DNA damage of up to 0.65 lesions/10�kb in the mouse, and 1.42 lesions /10�kb in the human. No significant effect on DNA damage was observed over this dose range in the nuclear DNA fragments investigated in these species. In contrast, tammar wallaby spermatozoa were susceptible to DNA damage at the 5�mM H2O2 dose in both nuclear (0.51 lesions/10�kb) and mitochondrial (0.55 lesions/10�kb) genomes. This study is the first to compare DNA damage in specific DNA sequences in spermatozoa of different mammalian species. Nuclear DNA of the metatherian species, the tammar wallaby, was more susceptible to oxidative damage than that of the eutherian species. A major difference between metatherian and eutherian spermatozoa is that, in general, the former possess protamines that are not stabilised by disulfide cross-linkage. These findings therefore suggest that sperm chromatin packaging affects the susceptibility of sperm DNA to oxidative damage. (1) Sawyer and Aitken (2000) Reprod. Med. Rev. 8, 107–126.

scholarly journals Loss of Prm1 leads to defective chromatin protamination, impaired PRM2 processing, reduced sperm motility and subfertility in male mice

2021 ◽  
Author(s):  
Gina Esther Merges ◽  
Julia Meier ◽  
Simon Schneider ◽  
Alexander Kruse ◽  
Andreas Christian Fröbius ◽  
...  
Keyword(s):  
Dna Damage ◽  
Gene Editing ◽  
Dna Integrity ◽  
General Stress Response ◽  
Nuclear Proteome ◽  
Overall Ratio ◽  
Species Specific ◽  
High Level ◽  
Key Events ◽  
Deficient Mice

One of the key events during spermiogenesis is the hypercondensation of chromatin by substitution of the majority of histones by protamines. In humans and mice, protamine 1 (PRM1/Prm1) and protamine 2 (PRM2/Prm2), are expressed in a species-specific ratio. Using CRISPR-Cas9-mediated gene editing we generated Prm1-deficient mice and demonstrate, that Prm1+/- mice are subfertile while Prm1-/- are infertile. Prm1-deficiency was associated with higher levels of 8-OHdG, an indicator for reactive oxygen mediated DNA-damage. While Prm1+/- males displayed moderate increased levels of 8-OHdG virtually all sperm of Prm1-/- males displayed ROS mediated DNA damage. Consequently, DNA integrity was slightly hampered in Prm1+/-, while DNA was completely fragmented in Prm1-/- animals. Interestingly CMA3 staining which indicates protamine-free DNA revealed, that Prm1+/- sperm displayed high levels (93%), compared to Prm2+/- (29%) and WT (2%) sperm. This is not due to increased histone retention as demonstrated by mass spectrometry (MassSpec) of nuclear proteins in Prm1+/- sperm. Further analysis of the MassSpec data from sperm nuclear proteome revealed, that only one protein (RPL31) is significantly higher abundant in Prm1+/- compared to WT sperm. Comparison of the proteome from Prm1-/- and Prm2-/- to WT suggested, that there are a small number of proteins which differ in abundance. However, their function was not linked mechanistically to primary defects seen in Prm1-/- mice and rather represent a general stress response. Interestingly, using acid urea gels we found that sperm from Prm1+/- and Prm1-/- mice contain a high level of unprocessed, full-length PRM2. Prm2 is transcribed as a precursor protein which, upon binding to DNA is successively processed. Further, the overall ratio of PRM1:PRM2 is skewed from 1:2 in WT to 1:5 in Prm1+/- animals. Our results reveal that Prm1 is required for proper processing of PRM2 to produce the mature PRM2 which, together with Prm1 is able to hypercondense DNA. Hence, the species specific PRM1:PRM2 ratio has to be precisely controlled in order to retain full fertility.

scholarly journals Species-Specific Proteins in the Oviducts of Snail Sibling Species: Proteotranscriptomic Study of Littorina fabalis and L. obtusata

Biology ◽  
2021 ◽  
Vol 10 (11) ◽  
pp. 1087
Author(s):  
Arseniy A. Lobov ◽  
Irina Y. Babkina ◽  
Lavrentii G. Danilov ◽  
Alexey E. Masharskiy ◽  
Alexander V. Predeus ◽  
...  
Keyword(s):  
Sibling Species ◽  
Reproductive System ◽  
Immune Defense ◽  
Illumina Hiseq ◽  
Littorina Obtusata ◽  
Associated Proteins ◽  
Specific Proteins ◽  
Species Specific ◽  
Molecular Patterns

Genus Littorina subgenus Neritrema (Mollusca, Caenogastropoda) includes the “obtusata” group of closely related species (Littorina obtusata and L. fabalis). The anatomy of the adult reproductive system (pallial oviduct) is the only reliable feature used for species identification in females of these species. Reproductive system anatomy and reproduction-associated proteins often diverge between sibling species. Despite being of high evolutionary interest, the molecular basis of this divergence remains poorly understood. We performed proteotranscriptomic comparison of oviducts of L. obtusata and L. fabalis by RNA-seq on Illumina HiSeq 2500 and two-dimensional protein electrophoresis (2D DIGE) with MS/MS identification of the species-specific proteins. The interspecies differences in the oviduct were associated with (1) metabolic proteins reflecting overall physiological differences between L. obtusata and L. fabalis, (2) receptor proteins, and (3) transcripts related to transposable elements (TEs). Various receptors identified may recognize a wide variety of ligands from pathogen-associated molecular patterns to specific carbohydrates on the sperm surface. Therefore, these may participate in immune defense as well as in sperm storage and regulation. Species-specificity of multiple TE sequences (coding for reverse transcriptase and ribonuclease H) may indicate the important role of these genomic elements in the Littorina species divergence, which has not been reported previously.

Sperm chromatin condensation and single‐ and double‐stranded DNA damage as important parameters to define male factor related recurrent miscarriage

2020 ◽  
Vol 87 (11) ◽  
pp. 1126-1132
Author(s):  
Jordi Ribas‐Maynou ◽  
Carlos Abad ◽  
Sergio García‐Segura ◽  
Maria Oliver‐Bonet ◽  
Elena Prada ◽  
...  
Keyword(s):  
Dna Damage ◽  
Recurrent Miscarriage ◽  
Chromatin Condensation ◽  
Sperm Chromatin ◽  
Male Factor ◽  
Double Stranded Dna ◽  
Sperm Chromatin Condensation

scholarly journals The role of the dehydration stage in the post-hypertonic hemolysis of mammalian erythrocytes

2021 ◽  
Vol 23 (2) ◽  
pp. 32-36
Author(s):  
O. E. Nipot ◽  
◽  
O. O. Shapkina ◽  
P. M. Zubov ◽  
N. V. Orlova ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Cell Division ◽  
Salt Concentration ◽  
Coefficient Of Variation ◽  
Mammalian Species ◽  
Rat Erythrocytes ◽  
Species Specific ◽  
Hypertonic Shock ◽  
Median Cell

The aim of this study was to assess the level of damage to mammalian erythrocytes under post-hypertonic shock depending on the concentration of NaCl in the dehydration medium and to determine the effect of hypertonic NaCl solutions on the condition of mammalian erythrocytes by flow cytometry. To achieve this goal, spectrophotometric and cytometry research methods were used. The data obtained showed that post-hypertonic lysis of mammalian erythrocytes depends on the concentration of NaCl in the dehydration medium. The most sensitive to the effects of post-hypertonic shock are rat erythrocytes, the least sensitive are rabbit cells. Cytometry studies revealed significant changes in the histograms of the distribution of erythrocytes of all mammalian species with increasing salt concentration in the dehydration medium. These changes are species-specific and are probably related to changes in cell volume and morphology. The data revealed a relationship between the level of post-hypertonic hemolysis and the values of such indicators as the median distribution and the coefficient of variation. Thus, an increase in the sensitivity of mammalian erythrocytes to post-hypertonic shock with increasing salt concentration in dehydration medium was usually accompanied by a decrease in the median cell division, and higher values of the coefficient of variation are characteristic of mammalian erythrocytes resistant to post-hypertonic shock.

scholarly journals Molecular morphology and function of bull spermatozoa linked to histones and associated with fertility

Reproduction ◽  
2013 ◽  
Vol 146 (3) ◽  
pp. 263-272 ◽  
Author(s):  
Rodrigo V de Oliveira ◽  
Sule Dogan ◽  
Lauren E Belser ◽  
Abdullah Kaya ◽  
Einko Topper ◽  
...  
Keyword(s):  
Chromatin Condensation ◽  
Mammalian Species ◽  
Sperm Concentration ◽  
Low Fertility ◽  
Sperm Chromatin ◽  
High Fertility ◽  
Expression Levels ◽  
Molecular Morphology ◽  
Sperm Chromatin Condensation

Sub-par fertility in bulls is influenced by alterations in sperm chromatin, and it might not be solved with increased sperm concentration in artificial insemination. Appropriate histone retention during sperm chromatin condensation plays critical roles in male fertility. The objective of this study was to determine failures of sperm chromatin condensation associated with abnormal persistence or accessibility of histones by aniline blue (ANBL) test, expression levels, and cellular localizations of one variant and two core histones (H3.3, H2B, and H4 respectively) in the spermatozoa of low-fertility (LF) vs high-fertility (HF) bulls. The expression levels and cellular localizations of histones in spermatozoa were studied using immunoblotting, immunocytochemistry, and staining methods. The bioinformatics focused on the sequence identity and evolutionary distance of these proteins among three mammalian species: bovine, mouse, and human. We demonstrated that ANBL staining was different within the LF (1.73 (0.55, 0.19)) and HF (0.67 (0.17, 0.06)) groups (P<0.0001), which was also negatively correlated within vivobull fertility (r=−0.90,P<0.0001). Although these histones were consistently detectable and specifically localized in bull sperm cells, they were not different between the two groups. Except H2B variants, H3.3 and H4 showed 100% identity and were evolutionarily conserved in bulls, mice and humans. The H2B variants were more conserved between bulls and humans, than in mice. In conclusion, we showed that H2B, H3.3, and H4 were detectable in bull spermatozoa and that sperm chromatin condensation status, changed by histone retention, is related to bull fertility.

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