Mice with a Targeted Disruption of the H1t Gene Are Fertile and Undergo Normal Changes in Structural Chromosomal Proteins During Spermiogenesis1

Fantz, Douglas A.; Hatfield, Wendy R.; Horvath, Gary C.; Kistler, Malathi K.; Kistler, W S

doi:10.1095/biolreprod64.2.425

Cited by 67 publications

(52 citation statements)

References 54 publications

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“…The task of its posttranslational modification may be to establish an open chromatin structure for the replacement of histones with transition proteins and protamines. However, the functional significance of H1t has been questioned, because mice lacking histone H1t are fertile and show normal spermatogenesis (62)(63)(64). Redundancy of the linker histones and compensation for H1t by other H1s may hide detection of defects caused by the loss of H1t.…”

Section: Samplementioning

confidence: 99%

Testis-specific Linker Histone H1t Is Multiply Phosphorylated during Spermatogenesis

Sarg

Chwatal

Talasz

et al. 2009

Journal of Biological Chemistry

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During normal spermatogenesis, the testis-specific linker histone H1t appears at pachytene stage becomes phosphorylated in early spermatids and disappears in late spermatids. Using reversed-phase and hydrophilic interaction liquid chromatography, H1t from rat and mouse testes was isolated, subjected to enzymatic digestion, and analyzed by mass spectrometry. We observed different phosphorylated states of H1t (mono-, di-, and triphosphorylated) as well as the unphosphorylated protein.Tandem mass spectrometry and immobilized metal ion affinity chromatography experiments with MS/MS/MS and multistage activation were utilized to identify five phosphorylation sites on H1t from rats. Phosphorylation occurs on both serine and threonine residues, whereas only two of these sites were located on peptides containing the CDK consensus motif (S/T)PXZ. Rat H1t phosphorylation starts first by phosphorylation of the nonconsensus motif SPKS in the COOH-terminal domain, namely at Ser-140 and to a smaller degree at a further nonconsensus motif at Ser-186. This is followed by phosphorylation of Ser-177 and Thr-155, both located in CDK consensus motifs. A single phosphorylation site at Ser-8 in the NH 2 -terminal tail was also found. Mouse H1t lacks Ser-186 and is phosphorylated at up to four sites. In contrast to somatic linker histones, no strict order of increasing phosphorylation could be detected in H1t. Thus, it appears that not the order of up-phosphorylation but the number of the phosphate groups is necessary for regulated chromatin decondensation, thus facilitating the substitution of H1t by transition proteins and protamines.Mammalian spermatogenesis is a complex, strictly organized process, including proliferation and differentiation. After several mitotic divisions of spermatogonia, germ cells differentiate into spermatocytes and pass through two consecutive meiotic divisions, resulting in haploid round cells termed spermatids. These spermatids then evolve into highly condensed and transcriptionally inert sperm through a process known as spermiogenesis (for a review, see Ref.

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Section: Samplementioning

confidence: 99%

Testis-specific Linker Histone H1t Is Multiply Phosphorylated during Spermatogenesis

Sarg

Chwatal

Talasz

et al. 2009

Journal of Biological Chemistry

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“…Each toroid contains 60 kb of DNA and is linked to other toroids by uncoiled DNA stretches. Whereas the roles of PRMs in sperm chromatin condensation and nuclear shaping are evident, the roles of H1t (16)(17)(18), TNP1 (20), and TNP2 (21) in the intermediate steps before the appearance of PRMs remain uncertain. Because HILS1 has diverged significantly from other linker histone family members and is expressed in elongating and elongated spermatids where core histones are absent, HILS1 is unlikely to function as a typical linker histone.…”

Section: Hils1 Is a Chromatin-associated Protein (Cp)mentioning

confidence: 99%

“…H1t is first detected in mid-pachytene spermatocytes, where it rapidly integrates into the chromatin, replaces most of the other somatic H1 subtypes, and persists until the elongating spermatid stage (15). Surprisingly, knockout of H1t does not affect chromatin remodeling, indicating that other known or unknown linker histones may compensate for its absence (16)(17)(18).The development of spermatids into spermatozoa, termed spermiogenesis, is characterized by striking morphological and molecular transformations. In elongating and elongated spermatids, major restructuring of the somatic chromatin occurs; this process involves the replacement of somatic histones and H1t by transition nuclear proteins (TNP1 and TNP2), which are subsequently replaced by protamines (PRM1 and PRM2).…”

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confidence: 99%

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confidence: 99%

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HILS1 is a spermatid-specific linker histone H1-like protein implicated in chromatin remodeling during mammalian spermiogenesis

Yan

Burns

et al. 2003

Proc. Natl. Acad. Sci. U.S.A.

165

136

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Chromatin remodeling is a major event that occurs during mammalian spermiogenesis, the process of spermatid maturation into spermatozoa. Nuclear condensation during spermiogenesis is accomplished by replacing somatic histones (linker histone H1 and core histones) and the testis-specific linker histone, H1t, with transition proteins and protamines. It has long been thought that H1t is the only testis-specific linker histone, and that all linker histones are replaced by transition proteins, and subsequently by protamines during spermiogenesis. Here, we report the identification and characterization of a spermatid-specific linker histone H1-like protein (termed HILS1) in the mouse and human. Both mouse and human HILS1 genes are located in intron 8 of the ␣-sarcoglycan genes. HILS1 is highly expressed in nuclei of elongating and elongated spermatids (steps 9 -15). HILS1 displays several biochemical properties that are similar to those of linker histones, including the abilities to bind reconstituted mononucleosomes, produce a chromatosome stop during micrococcal nuclease digestion, and aggregate chromatin. Because HILS1 is expressed in late spermatids that do not contain core histones, HILS1 may participate in spermatid nuclear condensation through a mechanism distinct from that of linker histones. Because HILS1 also belongs to the large winged helix͞forkhead protein superfamily, HILS1 may also regulate gene transcription, DNA repair, and͞or other chromosome processes during mammalian spermiogenesis.spermatogenesis ͉ transition nuclear proteins ͉ in silico subtraction ͉ genome database mining H istones are a family of basic proteins that organize eukaryotic DNA into a compact chromatin. There are five major classes of histones, the core histones H2A, H2B, H3, and H4, and the linker histones, H1. Two of each of the four core histones constitute an octamer unit of the nucleosome particle. H1 histones bind to DNA in the nucleosome and to the linker DNA between nucleosomes, thereby facilitating the compaction of nucleosomes into a 30-nm chromatin fiber and higher-order chromatin structures (1). These interactions between histones and DNA modulate gene activity, and both core and H1 histones have profound effects on transcription (2, 3). Among the five histone classes, H1 histones exhibit the most diversity. In mice and humans, there are eight previously described H1 subtypes, including the five somatic subtypes H1a-H1e, the replacement subtype H1o, the testis-specific linker-histone H1t (4-6), and the oocyte-specific H1 linker histone, H1foo (7). The genomic organizations of these histone H1 genes are conserved (8,9).Spermatogenesis is the process of development of male germ cells from spermatogonia to highly differentiated spermatozoa. During spermatogenesis, chromatin is restructured, and dramatic changes in histone gene expression are observed (10). In spermatogonia and preleptotene spermatocytes (cells undergoing DNA replication), the H1a and H1b linker histones are expressed (11). During meiotic prophase, striking ...

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“…In addition, male germ cells contain specialized chromatin components such as H1T, a histone H1 variant specifically expressed in pachytene spermatocytes and early haploid cells (4), and HILS1, a histone H1-related protein specifically expressed in elongating and elongated spermatids (5). However, no spermatogenesis abnormalities are observed in mice lacking the H1t gene, perhaps because of redundancy with the somatic H1.1, which is also expressed at this stage (6)(7)(8)(9). This finding contrasts with the reduced fertility and defective spermiogenesis in mice lacking transition proteins 1 or 2 or haploinsufficient for protamines 1 or 2 (10-12).…”

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Polar nuclear localization of H1T2, a histone H1 variant, required for spermatid elongation and DNA condensation during spermiogenesis

Martianov

Brancorsini

Catena

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

189

168

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Spermiogenesis entails a major biochemical and morphological restructuring of the germ cell involving replacement of the somatic histones by protamines packing the DNA into the condensed spermatid nucleus and elimination of the cytoplasm during the elongation phase. We describe H1T2, an histone H1 variant selectively and transiently expressed in male haploid germ cells during spermiogenesis. In round and elongating spermatids, H1T2 specifically localizes to a chromatin domain at the apical pole, revealing a polarity in the spermatid nucleus. Inactivation by homologous recombination shows that H1T2 is critical for spermiogenesis as male H1t2 ؊/؊ mice have greatly reduced fertility. Analysis of spermiogenesis in H1t2 mutant mice shows delayed nuclear condensation and aberrant elongation. As a result, mutant spermatids are characterized by the presence of residual cytoplasm, acrosome detachment, and fragmented DNA. Hence, H1T2 is a protein required for proper cell restructuring and DNA condensation during the elongation phase of spermiogenesis.chromatin ͉ mouse ͉ protamine ͉ acrosome M ammalian spermatogenesis involves the differentiation of diploid spermatogonia into spermatocytes and then, through two successive meiotic divisions, into haploid round spermatids. During spermiogenesis, the haploid round spermatids undergo an elongation phase, transforming them into mature spermatozoa. This process entails a major biochemical and morphological restructuring of the germ cell where the majority of the somatic histones are replaced, first by transition proteins, then protamines packing the DNA into the sperm cell nucleus.Differentiating male germ cells use specialized transcriptional regulatory mechanisms involving testis-specific paralogues of transcription factors such as TLF͞TRF2, ALF, and TAF7L (1-3). In addition, male germ cells contain specialized chromatin components such as H1T, a histone H1 variant specifically expressed in pachytene spermatocytes and early haploid cells (4), and HILS1, a histone H1-related protein specifically expressed in elongating and elongated spermatids (5). However, no spermatogenesis abnormalities are observed in mice lacking the H1t gene, perhaps because of redundancy with the somatic H1.1, which is also expressed at this stage (6-9). This finding contrasts with the reduced fertility and defective spermiogenesis in mice lacking transition proteins 1 or 2 or haploinsufficient for protamines 1 or 2 (10-12). Thus, whereas H1 histones have been proposed to play a role in the formation of higher-order chromatin structure, the physiological functions of the tissue-specific variants remain obscure. Here, we describe a male germ cellspecific H1-related protein, H1T2, that plays a critical role during the elongation phase of spermiogenesis. Materials and MethodsCloning and Inactivation of H1t2. The H1t2 cDNA was cloned from a mouse testis cDNA library by screening with the short mouse TEST640 EST probe as described (2, 13). 5Ј RACE using mouse testis RNA was performed to confirm the sequence o...

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Mice with a Targeted Disruption of the H1t Gene Are Fertile and Undergo Normal Changes in Structural Chromosomal Proteins During Spermiogenesis1

Cited by 67 publications

References 54 publications

Testis-specific Linker Histone H1t Is Multiply Phosphorylated during Spermatogenesis

Testis-specific Linker Histone H1t Is Multiply Phosphorylated during Spermatogenesis

HILS1 is a spermatid-specific linker histone H1-like protein implicated in chromatin remodeling during mammalian spermiogenesis

Polar nuclear localization of H1T2, a histone H1 variant, required for spermatid elongation and DNA condensation during spermiogenesis

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