Interaction of exogenous DNA with the nuclear matrix of live spermatozoa

McCarthy, Siobhán; Ward, W. Steven

doi:10.1002/(sici)1098-2795(200006)56:2+<235::aid-mrd4>3.0.co;2-d

Cited by 12 publications

(7 citation statements)

References 21 publications

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“…These results indicate that, regardless of the findings of a previous report [21] demonstrating interaction between exogenous DNA molecules and the nuclear matrix of mouse spermatozoa, the impact of transgene flanking with MARs on the efficiency of this procedure is not significant. Our results also suggest that the mechanism of transgene integration occurring after ICSI might not involve homologous recombination events mediated by MAR sequences.…”

Section: Discussioncontrasting

confidence: 49%

“…It has been recently reported that endogenous repeated sequences could be implicated in homologous recombination events in mouse embryos, involving 70% of integrated events in some experiments [20]. Also, the fact that live mouse spermatozoa can bind exogenous DNA molecules via interactions with the sperm nuclear matrix [21] further supports the importance of evaluating the impact of transgene flanking with MARs on the efficiency of ICSI-mediated transgenesis.…”

Section: Introductionmentioning

confidence: 96%

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Effect of Transgene Concentration, Flanking Matrix Attachment Regions, and RecA-Coating on the Efficiency of Mouse Transgenesis Mediated by Intracytoplasmic Sperm Injection1

Moreira

Pérez-Crespo

Ramírez

et al. 2007

Biology of Reproduction

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Intracytoplasmic sperm injection (ICSI) of DNA-loaded sperm cells has been shown to be a valuable tool for the production of transgenic animals, especially when DNA constructs with submegabase magnitude are used. In order to optimize and to understand the mechanism of the ICSI-mediated transgenesis, we have evaluated the impact of transgene DNA concentration, transgene flanking with nuclear matrix attachment regions (MARs), and the use of recombinase A (RecA)-coated DNA on the efficiency of mouse transgenesis production by ICSI. Presented data include assays with three DNA constructs; an enhanced green fluorescent protein (EGFP) plasmid of 5.4 kb, this plasmid flanked with two MAR elements (2.3 Kb of the human beta-interferon domain boundaries), and a yeast artificial chromosome (YAC) construct of approximately 510 kb (the largest transgenic construct introduced by ICSI that we have seen reported). ICSI-mediated transgenesis was done in the B6D2 mouse strain using different concentrations for each construct. Analysis of generated data indicated that ICSI allows the use of higher DNA concentrations than the ones used for pronuclear microinjection, however, when a certain threshold is exceeded, embryo/fetal viability decrease dramatically. In addition, independently of the transgene concentration tested, transgene flanking with MAR sequences did not have a significant impact on the efficiency of this transgenesis method. Finally, we observed that although the overall efficiency of ICSI-mediated transgenesis with fresh spermatozoa and RecA-complexed DNA was similar to the one obtained with the common ICSI-mediated transgenesis approach with frozen-thawed spermatozoa and RecA free DNA, this method was not as efficient in maintaining a low frequency of founder animal mosaicism, suggesting that different mechanisms of transgene integration might result from each procedure.

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Section: Discussioncontrasting

confidence: 49%

Section: Introductionmentioning

confidence: 96%

Effect of Transgene Concentration, Flanking Matrix Attachment Regions, and RecA-Coating on the Efficiency of Mouse Transgenesis Mediated by Intracytoplasmic Sperm Injection1

Moreira

Pérez-Crespo

Ramírez

et al. 2007

Biology of Reproduction

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“…In reference to the importance of chromatin structure, McCarthy and Ward (2000) described the sperm nucleus as an ''ordered library of DNA organized into functional zones.'' Histone-associated DNA in the male pronucleus is preferentially acetylated and transcriptionally active during S/G 2 (Adenot et al, 1997) and early embryonic development (GardinerGarden et al, 1998).…”

Section: Sperm Chromatin Structure In Embryonic Gene Expressionmentioning

confidence: 99%

Sperm Chromatin Structure Assay: Its Clinical Use for Detecting Sperm DNA Fragmentation in Male Infertility and Comparisons With Other Techniques

Evenson

Larson

Jost

2002

Journal of Andrology

909

768

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The Sperm Chromatin Structure Assay (SCSA) was first described in the December 5, 1980 issue of Science (Evenson et al, 1980a). The data described in that article showed a significant difference between proven fertile and subfertility or infertility in men and bulls and the susceptibility to denaturation of their sperm nuclear DNA. A subsequent study (Evenson et al, 1984) described SCSA data obtained from men with testicular cancer. The data were heterogeneous, and begged the question, what was SCSA measuring? Great effort has been spent over the past 15 years on animal model systems, mostly related to dose-response toxicology experiments and large animal fertility trials. The results from these studies showed that SCSA was highly dose-responsive to toxicants, highly repeatable, and provided meaningful biological information on sperm nuclear DNA defects. The extensive data from nonhumans (Evenson et al, 1985(Evenson et al, , 1989a(Evenson et al, ,b, 1993a(Evenson et al, ,b,c, 1994Ballachey et al, 1987Ballachey et al, , 1988Evenson and Jost, 1993; Sailer et al, 1995a,b;Evenson, 1999a), and human toxicology and fertility studies (Evenson et al, 1978(Evenson et al, , 1980a(Evenson et al, ,b, 1984(Evenson et al, , 1991Evenson and Melamed, 1983;Evenson, 1999b;Larson et al, 1999Larson et al, , 2000Larson et al, , 2001 provide This work was supported in part by grant R827019 from the US Environmental Protection Agency, grants EPS-9720642 and OSR-9452894 from the National Science Foundation, and by SCSA Diagnostics, Inc. This is South Dakota Agricultural Experiment Station publication 3273 of the journal series.Correspondence to: Donald P. Evenson, Olson Biochemistry Laboratories, ASC 136, South Dakota State University, Brookings, SD 57006 (e-mail: donald evenson@sdstate.edu).Received for publication September 20, 2001; accepted for publication September 20, 2001. compelling evidence that SCSA will be useful in clinical human semen analysis (Figure 1).After reviewing current infertility center sites on the World Wide Web and approximately a dozen lay-oriented books on infertility and assisted reproductive technology (ART), it was amazing that not a single reference contained any information about the negative influence of fragmented sperm nuclear DNA on successful pregnancy outcome. Furthermore, few physicians are aware of scientific literature on this subject. Currently, if a spermatozoon has some motility as seen under a light microscope, and reasonable morphology, then many ART clinics assume that the sperm DNA must be fine. However, a number of cases have been documented in which couples have had multiple failed attempts at intracytoplasmic sperm injection (ICSI), even using donor eggs, and we found that the fragmentation of sperm DNA was above our threshold for less than 1% probability of achieving a successful pregnancy. Spermatozoa with defective DNA can fertilize an oocyte, produce high-quality early stage embryos, and then, in relationship to the extent of DNA damage, fail in producing a successful term ...

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“…For instance, several studies have shown that spermatozoa integrate DNA at one, or a few, specific site(s), and multiple instances of integration at the same site were observed. [90][91][92] The authors suggested that there may be relatively few sites accessible for DNA integration, as the probability of DNA integrating in the same locus in two separate clones should be extremely low. In support of this, a preintegrated locus was shown to be 75-to 470-fold more likely to receive another incoming vector than any randomly assigned locus.…”

Section: Illegitimate Integration Sites Are Not Totally Randommentioning

confidence: 99%

Illegitimate DNA integration in mammalian cells

2003

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Foreign DNA integration is one of the most widely exploited cellular processes in molecular biology. Its technical use permits us to alter a cellular genome by incorporating a fragment of foreign DNA into the chromosomal DNA. This process employs the cell's own endogenous DNA modification and repair machinery. Two main classes of integration mechanisms exist: those that draw on sequence similarity between the foreign and genomic sequences to carry out homology-directed modifications, and the nonhomologous or 'illegitimate' insertion of foreign DNA into the genome. Gene therapy procedures can result in illegitimate integration of introduced sequences and thus pose a risk of unforeseeable genomic alterations. The choice of insertion site, the degree to which the foreign DNA and endogenous locus are modified before or during integration, and the resulting impact on structure, expression, and stability of the genome are all factors of illegitimate DNA integration that must be considered, in particular when designing genetic therapies.

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Interaction of exogenous DNA with the nuclear matrix of live spermatozoa

Cited by 12 publications

References 21 publications

Effect of Transgene Concentration, Flanking Matrix Attachment Regions, and RecA-Coating on the Efficiency of Mouse Transgenesis Mediated by Intracytoplasmic Sperm Injection1

Effect of Transgene Concentration, Flanking Matrix Attachment Regions, and RecA-Coating on the Efficiency of Mouse Transgenesis Mediated by Intracytoplasmic Sperm Injection1

Sperm Chromatin Structure Assay: Its Clinical Use for Detecting Sperm DNA Fragmentation in Male Infertility and Comparisons With Other Techniques

Illegitimate DNA integration in mammalian cells

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