DNA Unpacking in Guinea Pig Sperm Chromatin by Heparin and Reduced Glutathione

Abstract

The kinetics of nuclear decondensation and DNA unpacking induced by the action of a physiological concentration of heparin and glutathione of guinea pig spermatozoa was studied. Sperm (acrosomeless) suspensions were incubated at several different temperatures (37, 40, 43, and 46 degrees C), with a constant concentration of either heparin (50 microM) or reduced glutathione (12.5 mM) and increasing concentrations of the other reagent. Nuclei spermatozoa remained highly condensed when incubated in the medium alone or in either GSH or heparin alone for up to 72 h. Swelling of nuclei spermatozoa was initially observed during the first 20 min of incubation. The sperm nuclei initiate decompaction at the central part of the nuclear structure while at the periphery there remain numerous residues of densely packed chromatin. The swollen chromatin pattern presents the characteristic organization into "hub-like" nuclear bodies that measured 10-100 nm diameter joined by a network of chromatin fibers. At full nuclei decondensation chromatin end fibers are loose, probably meaning that DNA is not organized into loop domains. DNA presence was verified by the use of ethidium bromide and acridine orange.

Chromosome architecture in the decondensing human sperm nucleus

“…The chromatin beads of 500 nm observed by us might correspond to the 600 nm units, although it is difficult to compare data because of fundamentally different approaches of sample preparation. Combinations of nodular (beaded) and fiber (cordlike) structures in sperm chromatin were reported earlier: 'knobby' fibers 33-42 nm and 65-120 nm width laced together by fine 6-8 nm strands (Sobhon et al, 1982); 'knobby' fibers 11 nm width (Fraschini and Biggiogera, 1985); 'hub-like' nuclear bodies with diameter of 10-100 nm, joined by a network of chromatin fibers (Sanchez-Vazquez et al, 1998); globular structures of 120-150 Å in diameter (Gusse and Chevaillier, 1980). These EM studies, while providing more accurate distance measurements, did not distinguish between chromatin belonging to different chromosomes and chromosome arms, whereas our approach, based on light microscopy, cannot provide structural data at distances less then 300 nm.…”

“…On the higher structural level, it has been proposed that chromatin in mammalian sperm is organized into loop domains attached at their bases to a nuclear matrix (Ward and Coffey, 1991;Yaron et al, 1998;Ward and Ward, 2004). Looped organization of sperm DNA is disputed in other works (Sanchez-Vazquez et al, 1998).…”

Spermatozoal transcriptome profiling for bull sperm motility: a potential tool to evaluate semen quality

“…On the higher structural level, it has been proposed that chromatin in mammalian sperm is organized into loop domains attached at their bases to a nuclear matrix (Ward & Coffey 1991, Yaron et al 1998, Ward & Ward 2004. Looped organization of sperm DNA is disputed in other works (Sanchez-Vazquez et al 1998) but the presence and localization of specific RNA within or close to the sperm nuclear matrix have recently been demonstrated (Lalancette et al 2008a). It is thus logical to speculate that a more drastic hot-TRIzol protocol was required to show up transcripts enclosed into more complex and less accessible chromatin organization.…”

Rapid coupling between gravitational forces and the transcriptome in human myelomonocytic U937 cells

“…Therefore, the preceding hypothetical transduction processes have to be fast enough for inducing transcriptome changes within 20 s of force alteration. We therefore compiled the transduction velocities of potential transduction processes: DNA decondensation was shown to occur within the time frame of minutes to hours 66 , transcription factor binding takes a couple of seconds 67 , and the elongation rate of RNA polymerase II was found to range between 0.37 kb/min 68 and 4.3 kb/min 69 with a median rate of 2.1 kb/min 68 . Theoretical velocities even above 50 kb/min have been calculated 70 .…”