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Hum. Reprod. Advance Access published online on November 16, 2006
Human Reproduction, doi:10.1093/humrep/del439
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© The Author 2006. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oxfordjournals.org
Received August 8, 2006
Revised October 3, 2006
Accepted October 13, 2006

Article

Testis/sperm-specific histone 2B in the sperm of donors and subfertile patients: variability and relation to chromatin packaging

S. Singleton 1, A. Zalensky 1, G.F. Doncel 2, M. Morshedi 1, and I.A. Zalenskaya 2 *

1 The Jones Institute for Reproductive Medicine, Eastern Virginia Medical School, Norfolk VA, USA
2 CONRAD, Department of Obstetrics and Gynecology, Eastern Virginia Medical School, Norfolk VA, USA

* To whom correspondence should be addressed.
I.A. Zalenskaya, E-mail: zalensia{at}evms.edu


  Abstract

BACKGROUND: The compaction of human sperm chromatin is the result of replacement of ~85% of histones with protamines. Germ-line testis/sperm-specific histone 2B (TSH2B) has been detected in only ~30% of mature spermatozoa. Its level in the semen of subfertile patients varies; its function is unknown. We evaluated TSH2B in the sperm samples of 23 donors and 49 subfertile patients and assessed its association with chromatin compaction status. METHODS: TSH2B level was measured using immunoblotting. Chromatin packaging quality was evaluated by staining with chromomycin A3 (CMA3) which marked spermatozoa with defective packaging. To assess both TSH2B and chromatin status in the same spermatozoon, CMA3 staining and TSH2B immunolocalization were performed sequentially. RESULTS: A significant correlation (r = 0.55, P = 0.0027) was found between TSH2B level and percentage of CMA3-positive sperm in patient and donor semen samples. When individual spermatozoa were assessed for these parameters, 92% of TSH2B-containing cells were also CMA3 positive. Variation in the total sperm TSH2B level was less in donors than in patients. CONCLUSIONS: CMA3 positive staining of TSH2B-containing individual spermatozoa and a significant correlation between the total TSH2B level and CMA3 percentage in semen samples suggest a structural role for TSH2B in sperm chromatin organization. Low variability of TSH2B level in donors implies a mechanism (however unknown) regulating this parameter.

Keywords: chromatin/chromomycin A3/histone TSH2B/male infertility/sperm.
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