Histone 1 Research Articles

Comparative analysis of testis transcriptomes associated with male infertility in triploid cyprinid fish.

Spermatogenesis involves a series of cellular transformations and thousands of regulated genes. Previously, we showed that the triploid fish (3nBY) cannot produce mature spermatozoa. In the present study, evaluation of the testis microstructure revealed that germ cells of 3nBY could develop into round spermatids, but then degenerated, resulting in male infertility. In this study we comparatively analysed the testis transcriptomes from 3nBY and its diploid parent YB and identified a series of differentially expressed genes (DEGs) that were enriched in the Wnt signalling pathway and the apoptotic and ubiquitin-mediated proteolysis processes in 3nBY. Gene ontology functional analyses revealed that some DEGs in 3nBY were directly associated with the process of gamete generation, development and sperm flagellum assembly. In addition, the expression of a number of genes related to meiosis (Inhibitor Of DNA Binding 2 (ID2), Ovo Like Transcriptional Repressor 1 (OVOL1)), mitochondria (ATP1b (ATPase Na+/K+ Transporting Subunit Beta 1), ATP2a (ATPase, Ca++ Transporting, Cardiac Muscle, Slow Twitch 2), ATP5a (ATP Synthase F1 Subunit Alpha), Mitochondrially Encoded Cytochrome C Oxidase I (COX1), NADH Dehydrogenase Subunit 4 (ND4)) and chromatin structure (Histone 1 (H1), Histone 2a (H2A), Histone 2b (H2B), Histone 3 (H3), Histone 4 (H4)) was lower in the testes of 3nBY, whereas the expression of genes encoding ubiquitin (Ubiquitin Conjugating Enzymes (UBEs), Ring Finger Proteins (RNFs)) and apoptosis (CASPs (Caspase 3, Caspase 7,Caspase 8), BCLs (B-Cell Lymphoma 3, B-Cell CLL/Lymphoma 2, B Cell CLL/Lymphoma 10)) proteins involved in spermatid degeneration was higher. These data suggest that the disrupted expression of genes associated with spermatogenesis and the increased expression of mitochondrial ubiquitin, which initiates cell apoptosis, may result in spermatid degeneration in male 3nBY. This study provides information regarding the potential molecular regulatory mechanisms underlying male infertility in polyploid fish.

Pullulan-PEI-Histidine towards gene delivery: vector unpacking with respect to molecular weight and cytoplasmic histones

Event Abstract Back to Event Pullulan-PEI-Histidine towards gene delivery: vector unpacking with respect to molecular weight and cytoplasmic histones Rekha M. Ramesan1 1 Sree Chitra Tirunal Institute for Medical Sciences & Technology, Biosurface Technology, BiomedicalTechnology Wing, India Introduction: Cationic polymers, one of the most important nonviral vectors have a number of advantages over viral vectors. However cationic surface charge, the key factor which determines the transfection efficiency, is also one of the limitations associated with it. Major limiting factors are cytotoxicity, biocompatibility and the release of the cargo from the vector intracellularly. The tightly formed nanoplexes gains entry into the cells, however it doesn’t lead to good transfection if the DNA remains tightly bound to carrier. Schaffer et.al proved that unpacking of the DNA from the vectors is a major limiting step in the process of non-viral gene delivery[1]. Earlier we reported the possible role of histones in unpacking in which a combination of all histones were used[2]. In this work we attempted to evaluate the possible role played by the molecular weight of the polymers and which of the cytoplasmic histones aids more in vector unpacking. Histone 1 (H1) and histone 3 (H3) reported to be found in cytoplasm were used for the study. Materials and Methods: Pullulan a hydrophilic, nonionic polysaccharide was conjugated with PEI (10 and 25KDa) to which histidine was introduced to improve its transfection efficiency (PPE10H and PPE25H). The conjugation was extablished by variuos techniques including NMR. The particle size and zetapotential of nanoplexes was evaluated with DLS. The cellular uptake and transfection efficiency was done in C6 glioma cells. For vector unpacking studies, cellulose-DNA beads were used by which the displacement of the polymer was assessed. Labelled PPEH and histones (H 1 and H 3) were used to study the unpacking in cell lines. Results and Discussion: The derivatisation was established using techniques including NMR and Raman spectroscopy. The optimum size of PPE10H and PPE25H nanoplexes were 78.4 ± 0.72 and 101.6 ± 2.3 respectively at 1:4 ratio with a zeta potential in the range 13-15 mV. The nanoplexes of both polymers demonstrated very high intracellular uptake and PPE10H had higher transfection efficiency in comparison with PEI10K and PPE10KH. Among the histones studied H3 was found to be more efficient in vector unpacking (Fig.1). In the absence of histone intact nanoplex is internalised as TRITC labeled PPE10H is found in the cytoplasm and tagged DNA in the nucleus. The nanoplex treated with H1 and H3 resulted in unpacking outside the cell and the labeled polymers are excluded from the entry into the cells (Fig. 1 B to E). However H1 is not as efficient as H3 in unpacking the DNA from the polyplex. The vector unpacking was higher with PPEH10K which might be the reason for better transfection efficacy which could be attributed to its low cationicity (Fig. 2). Figure 1. Internalization of TRITC labeled PPE10KH nanoplex in C6 cells. Nanoplex alone (A), nanoplex treated with 500 ng histone H1 (B), 1 micro g histone H1 (C), 500 ng histone H3 (D) and 1 micro g histone H3 (E) . Absence of TRITC labeled polymer in D & E indicate the better displacement of polymer with H3. Figure 2. Luciferase expression in C6 cells transfected by PPE10H and PEI complexes at ratios 1:3 and 1:4. ** p<0.05 Conclusion: PPE10H was found to be a better transfecting agent. It was observed that H3 is more effiecient in displacing the plasmid DNA from the vector. The extent of cationicity also found to play a role in release of DNA from the vector in presence of histones.

Overexpression of histone H2A modulates drug susceptibility in Leishmania parasites

Resistance to antimonials has emerged as a major hurdle to the treatment and control of visceral leishmaniasis (VL), also know as kala-azar (KA), the disease caused by Leishmania donovani, in India where >60% of KA patients are unresponsive to sodium antimony gluconate (SAG) treatment. Determinants of resistance in laboratory strains are partly known, however the mechanism operating in field isolates is not well understood. In microarray-based expression profiling with RNA isolated from field isolates of drug-resistant and -sensitive L. donovani parasites, genes encoding histone 1 (H1), histone 2A (H2A), histone 4 (H4), mitogen-activated protein kinase 1 (MAPK1) and two hypothetical proteins showed significantly higher expression in antimony-resistant parasites, whilst genes encoding an amino acid transporter showed higher expression in sensitive parasites. The expression level of these genes was validated by semiquantitative polymerase chain reaction (PCR). Furthermore, the higher expression of H1, H2A and MAPK1 was confirmed at the protein level in resistant isolates. Overexpression of H2A in a drug-sensitive laboratory strain as well as a field isolate of L. donovani resulted in conversion of SAG-sensitive Leishmania parasites into a resistant phenotype. Moreover, H2A overexpression resulted in a significant decrease in susceptibility towards other antileishmanial drugs currently in use, i.e. amphotericin B and miltefosine, pointing to its role in drug resistance.

Sequence Dependent Femtosecond-Resolved Hydration Dynamics in the Minor Groove of DNA and Histone—DNA Complexes

Understanding the sequence dependent molecular recognition of DNA is crucial for the rational design of many drugs. Femtosecond resolved studies on the hydration dynamics of the dodecamer duplexes having sequences (CGCGAATTCGCG)2 and (CGCAAATTTGCG)2, and their complexes with the nucleic protein histone 1 (H1) reveal significant correlation of the molecular recognition of the DNA and DNA-protein complexes with the dynamics of hydration. The different molecular recognition of DNA and DNA-protein complexes is also borne out by circular dichroism (CD) and fluorescence detected CD measurements.

Expression of histone 1 (H1) and testis-specific histone 1 (H1t) genes during stallion spermatogenesis

In eukaryotic cells, the major protein constituents of the chromatin are histones, which can be divided into five classes, identified as H1, H2A, H2B, H3 and H4. During normal spermatogenesis, a testis-specific H1t is expressed in primary spermatocytes and believed to facilitate histone to protamine exchanges during spermiogenesis. In equine testes we detected the H1 protein at 22 kDa by western blot analysis while H1t was detected at 29 kDa. H1 protein was found to be expressed in all germ cells up to elongating spermatids (Sc) at stage IV. In peripubertal animals, there was a prolonged expression up to elongating spermatids (Sd1) at stage V. A fragment of the equine H1t gene was cloned (GenBank Accession No. AJ865320). The mRNA expression of H1t was found at the level in spermatogonia and in primary spermatocytes up to mid-pachytene at stage VIII/I, whereas H1t protein was found to be expressed up to round spermatides (Sa/Sb1) at stage VIII/I. In peripubertal animals, the H1t protein expression was detected up to elongating spermatids (Sb2) at stage II. Analysis of testes of different ages (≤2 years) and (≥3 years) by real-time RT-PCR revealed an increase of H1t mRNA expression, with a wide range of individual variety between 2 and 4 years old animals indicating a stable expression in animals older than 4 years old. This is the first study to show the testis-specific H1t in the stallion and gives evidence that the well-known peripubertal infertility in the stallion may be related to an insufficient histone to protamine exchange. The pattern of protamine gene expression, however, has still to be elucidated.

ECRG2 Disruption Leads to Centrosome Amplification and Spindle Checkpoint Defects Contributing Chromosome Instability

Cancer cells contain an abnormal number of chromosomes (aneuploidy), which is a prevalent form of genetic instability in human cancers. Abnormal amplification of centrosomes and defects of spindle assembly checkpoint are the major causes of chromosome instability in cancer cells. Here we present biochemical evidence to suggest a role of ECRG2, a novel tumor suppressor gene, in maintaining chromosome stability. ECRG2 localized to centrosomes during interphase and kinetochores during mitosis. Further analysis revealed that ECRG2 participates in centrosome amplification in a p53-dependent manner. Depletion of ECRG2 not only destabilized p53, down-regulated p21, and increased the cyclin E/CDK2 activity, thus initiating centrosome amplification, but also abolished the ability of p53 localize to centrosomes. Overexpression of ECRG2 restored the p53-dependent suppression of centrosome duplication. Furthermore, ECRG2-depleted cells show severely disrupted spindle phenotype but fail to maintain the mitotic arrest due to minimal BUBR1 protein levels. Taken together, our results indicate that ECRG2 is important for ensuring centrosome duplication, spindle assembly checkpoint, and accurate chromosome segregation, and its depletion may contribute to chromosome instability and aneuploidy in human cancers.

Analysis of selected transcript levels in porcine spermatozoa, oocytes, zygotes and two-cell stage embryos

It has been suggested that spermatozoa can deliver mRNAs to the oocyte during fertilisation. Using reverse transcription and real-time quantitative polymerase chain reaction analysis (RQ-PCR), we evaluated the presence of clusterin (CLU), protamine 2 (PRM2), calmegin (CLGN), cAMP-response element modulator protein (CREM), methyltransferase 1 (DNMT1), linker histone 1 (H1), protamine 1 (PRM1), TATA box-binding protein associated factor 1 (TAF1) and TATA box-binding protein (TBP) in porcine mature oocytes, zygotes and two-cell stage embryos. Spermatozoa isolated from semen samples of boars contained all transcripts investigated, whereas oocytes contained only CREM, H1, TAF1, and TBP mRNAs. The zygote and two-cell stage embryos contained CLU, CREM, H1, PRM1, PRM2, TAF1 and TBP transcripts. Our observations suggest that porcine spermatozoa may delivery CLU, PRM1 and PRM2 mRNAs to the oocyte, which may contribute to zygotic and early embryonic development.

Interaction of Hoechst 33258 and Ethidium with Histone1−DNA Condensates

We report structural and dynamical aspects of DNAs from various sources including synthetic oligonucleotides in bulk buffer and as a complex with histone1 (H1). High-resolution transmission electron microscopic (HRTEM) studies reveal the structural change of the DNAs upon complexation with H1 leading to formation of compact-globular and hollow-toroidal particles. In order to explore the functionality of ligand binding of the DNAs and their complexes with H1, we have used two biologically common fluorescent probes Hoechst 33258 (H33258) and Ethidium (EB) as model ligands. Picosecond resolved fluorescence and polarization gated anisotropy studies examined that the minor groove binding of H33258 to the genomic DNA-H1 complex remains almost unaltered. However, the intercalative interaction of EB with the DNA in the complex is severely perturbed compared to that with the DNA in bulk buffer. Time-dependent solvochromic effect of the probe H33258 further elucidates the dynamical solvation, which is reflective of the overall environmental relaxation of the DNAs upon condensation by H1. We have also performed circular dichroism (CD) studies on the DNAs and their complexes with H1, which reveal the change in conformation of the DNAs in the complexes. Our studies in the ligand-binding mechanisms of the DNA-H1 complex are important to understand the mechanism of drug binding to linker DNA in condensed chromatin.

Immunization with H1, HASPB1 and MML Leishmania proteins in a vaccine trial against experimental canine leishmaniasis

The protective capabilities of three Leishmania recombinant proteins – histone 1 (H1) and hydrophilic acylated surface protein B1 (HASPB1) immunized singly, or together as a protein cocktail vaccine with Montanide™, and the polyprotein MML immunized with MPL®-SE adjuvant – were assessed in beagle dogs. Clinical examination of the dogs was carried out periodically under blinded conditions and the condition of the dogs defined as asymptomatic or symptomatic. At the end of the trial, we were able to confirm that following infection with L. infantum promastigotes, five out of eight dogs immunized with H1 Montanide™, and four out of eight dogs immunized with either the combination of HASPB1 with Montanide™ or the combination of H1 + HASPB1 with Montanide™, remained free of clinical signs, compared with two out of seven dogs immunized with the polyprotein MML and adjuvant MPL®-SE, and two out of eight dogs in the control group. The results demonstrate that HASPB1 and H1 antigens in combination with Montanide™ were able to induce partial protection against canine leishmaniasis, even under extreme experimental challenge conditions.

Chemical agents that promote chromatin compaction radiosensitize tumour cells

Purpose : Previous studies indicated that cells whose chromatin is naturally compacted at the time of radiation are hypersensitive to radiation-induced killing, primarily by single-hit inactivation. Some chemicals that are known to promote chromatin compaction in interphase cells are here investigated for their radiosensitizing potential. Materials and methods : Okadaic acid (OA), a protein phosphatase inhibitor, fostriecin (FC), a topoisomerase II inhibitor and trichostatin A (TSA), a histone deacetylase inhibitor, were reported to promote chromatin compaction in mammalian cells. Asynchronous populations of HT-29 (human colon carcinoma) cells were exposed to various concentrations of OA, FC and TSA for various times before irradiation with various doses of Cs-137 n -rays and toxicity and radiosensitization were measured. Induced chromatin compaction was visualized by electron microscopy (EM). Histone 1 (H1) and histone 3 (H3) phosphorylation was measured by Western blotting, whole-cell fluorescence microscopy and confocal microscopy. Results : OA and FC produced significant radiosensitization at 2 Gy after short (2 h) exposures. These chemical treatments also produced increased phosphorylation of H3 and increased chromatin compaction as measured by EM. A 2-h exposure of cells to TSA had no effect on cell radiosensitivity, histone phosphorylation or chromatin condensation. However, a 16-h exposure to TSA produced significant radiosensitization, histone phosphorylation and chromatin condensation, presumably by secondary mechanisms. Conclusions : These data are consistent with the hypothesis that compacted chromatin is a hypersensitive target for radiation killing. Furthermore, the modulation of chromatin conformation by drugs selectively in tumour cells might radiosensitize tumours whose cells are intrinsically radioresistant.

Histone Adduction with Nicotine: A Bio-Ams Study

Based on the study of DNA adduction with nicotine, we have measured the mouse hepatic histone adduction with14C-labeled nicotinein vivoby bio-accelerator mass spectrometry (bio-AMS). In the exposure of mice to nicotine, the dose range administered was from 0.2 μg to 6.0 μg kg b.w.-1, which was equivalent to a very low level of human exposure to cigarette smoke. The adducts of either histone 1 (H1) or histone 3 (H3) with nicotine in mouse liver increased markedly with increasing nicotine dose. Our results have demonstrated that in the study of protein adduction with toxic xenobiotics as a biomarker, the AMS method achieves the highest sensitivity, 4.6 × 10-17mol (46 amol) adducts per mg H1 protein, compared to all the other methods used previously.

Antibodies from patients with systemic lupus erythematosus and drug-induced lupus bind determinants on histone 5 (H5)

The antigenic domains of histone 5 (H5), a highly conserved variant of histone 1 (H1), were studied in relation to their reactivity with autoantibodies found in the sera of patients with systemic lupus erythematosus (SLE) and drug-induced lupus (DIL). While some H5 antibodies cross-react with H1, adsorption and immunoblotting studies have identified H5-specific antibodies as well. After proteolytic cleavage of H5 peptides, the reactivity of sera from these patients was tested by Western immunoblotting. All SLE ( 9 9 ) and DIL ( 7 7 ) sera bound an antigenic determinant in the carboxyl (C) terminus of H5 while none of the sera bound to the amino (N) terminus or the central hydrophobic domain. Although the reactivity of DIL sera with the purified H5 peptides was weaker than that of SLE sera, the antigenic domains bound by both groups of sera were the same. These observations demonstrate that the H5 domains reacting with DIL sera are restricted to the carboxyl terminus and are therefore no less restricted than those reacting with SLE sera. Further, the potential epitopes in the carboxyl terminus of H5 do not have a high degree of sequence identity with known mammalian peptides.

Purification of histone H1 polypeptides by high-performance cation-exchange chromatography

Calf thymus histone 1 (H1) was cleaved by chemical and enzymatic methods and the resulting polypeptides were fractionated by high-performance cation-exchange. Up to 1 mg of H1 polypeptides were loaded onto a 50 × 5 mm I.D. cation-exchange column and fractionated to greater than 95% purity in less than 30 min. This is the first report on the separation of H1 polypeptides by a strong cation-exchange matrix. In addition, the high-performance cation-exchange chromatography protocol represents a significant decrease in fractionation time when compared to conventional ion-exchange and gel filtration chromatography. The utility of this procedure is shown when the H1 peptides purified by the protocol were used to define antigenic domains of H1 band by procainamide-induced lupus and idiopathic systemic lupus erythematosus. The majority of the sera tested by enzyme-linked immunoassay (ELISA) reacted to the C-terminal peptides of H1 indicating this to be the major antigenic domain of H1.

A histone 1-like antigen is a component of the nuclear envelope

Rabbit antibodies have been raised against rat liver nuclear envelopes. An enzyme-linked immunosorbent assay (ELISA) demonstrated high titer antiserum specific for the nuclear envelope preparation. Immunocytochemical studies showed that the antiserum stained the nuclear envelopes, but not intra-nuclear components of HEp-2 (human malignant epithelial) cells. When electrophoretically separated peptides were tested by immunoblotting techniques, the rabbit antiserum specifically stained proteins with molecular masses of 26 and 28 kD. These peptides had similar mobilities to purified histone 1 (H1). Indeed purified calf thymus H1 recognized the antiserum. The antigens are not loosely bound to the nuclear envelope, as they could not be extracted with low salt. Therefore, we have established that the 26 and 28 kD nuclear envelope peptides are not contaminants of the nuclear envelope preparation and that they express determinants that are immunologically cross-reactive with purified H1, but not with intra-nuclear H1.

Antibodies in procainamide-induced and systemic lupus erythematosus bind the C-terminus of histone 1 (H1)

The sera of patients with systemic lupus erythematosus (SLE) and drug-induced lupus (DIL) were used to study the antigenic regions of histone 1 (H1) that bind antibodies in these sera. ELISA and immunoblotting techniques using enzymatically and chemically derived peptides of H1 showed that the major antigenic domain is in the carboxyl ( C) terminus. None of the 24 SLE or 11 DIL sera bound to the central hydrophobic polypeptide by ELISA. The reactivity of DIL sera with the purified H1 peptides was similar to that observed with SLE sera. This observation suggests a common immune pathway for DIL and SLE.

Distribution and synthesis of histone 1 subfractions during spermatogenesis in the rat

The distribution and synthesis of various histone 1 (H1) subfractions have been studied in purified populations of rat testis germinal cells. Rat testis H1 has been resolved by SDS polyacrylamide gel electrophoresis into four bands containing H1 subfractions H1b + H1d + H1e (H1bde), H1a, H1c, and H1t. Results show that the somatic H1 subfractions H1a, b, c, d, and e are present and synthesized in spermatogonia and early primary spermatocytes. H1a, a minor somatic subfraction, is the predominant H1 subfraction in these cells (45% of total H1). The testis-specific H1 subfraction, H1t, is barely detectable in these cells. In pachytene spermatocytes, however, H1t (56% of total H1) has replaced H1b, c, d and e, and levels of H1a remain high (30% of total H1). H1t is the only subfraction actively synthesized in pachytene spermatocytes. Early spermatids (steps 1–8) have the same subfraction composition as pachytene spermatocytes, but extremely low H1 synthesis occurs. These results show that gene expression of H1 subfractions is under developmental control. There appears to be an elevated synthesis of subfraction H1a associated with DNA replication. The localization of H1t synthesis to the pachytene spermatocytes corresponds to the regulation of testis-specific variants of H2B and H2A.

Lysines of histone 1 represent the principal target for covalent binding of microsomally activated benzo[a]pyrene in vitro.

The principal target amino acid residue for covalent binding of metabolically activated [3H]benzo[a]pyrene (B[a]P) to histone 1 (H1) has been identified. Highly purified calf thymus H1 was modified by incubation with [3H]B[a]P in the presence of rat liver microsomes. The relative distribution of [3H]B[a]P versus specific amino acids among the N-bromo-succinimide peptides of H1 suggested that lysine may be the target. This was tested by selectively blocking the nucleophilic amino groups of lysine by amidination prior to [3H]B[a]P binding. Increasing levels of amidination of H1 resulted in corresponding decreases in the reactivity of this histone toward [3H]B[a]P. Amidination of 93% of the lysines blocked 87% of the [3H]B[a]P binding indicating that lysine was the principal target. N-terminal amino acid sequencing of [3H]B[a]P-modified H1 confirmed that lysine residues had bound [3H]B[a]P.

Correlation of chromatin composition with metabolic changes in nuclei of primitive erythroid cells from chicken embryos.

Changes in levels of biosynthesis of DNA, RNA, and histones were compared with relative proportions of each histone class during primitive erythropoiesis in embryonic chicks. We confirmed that erythrocyte-specific histone 5 (H5) was substantial in the earliest accessible, erythroblast-enriched stage and that it doubled in relative amount between polychromatic and orthochromatic stages to about 1 mol per 2 mol of each nucleosomal histone, still considerable less than in adult definitive erythrocytes. No other histones changed during primitive erythropoiesis, but the molar proportion of histone 1 (H1) always exceeded that of H5 in these cells, unlike definitive erythrocytes. The increase in content of H5 was accompanied by continued decline in synthesis of the other histones and DNA. The accumulation of H5 during development appears to occur in steps corresponding to the maturation of the primitive and definitive erythroid cell lines. Lysine-rich histones were more easily extracted from nuclei of the erythrosynthesis in whole cells and in isolated nuclei.

Chromatin proteins as a possible target for antitumour agents: Alterations of chromatin proteins in dibromodulcitol treated Yoshida tumors

The effect of dibromodulcitol (DBD) on Yoshida sarcoma chromatin components has been investigated. Measurements on the radioactivity of nuclear components after in vivo treatment with [3H]DBD for 1 h indicated preferential drug binding to the high molecular weight component of the nuclear residual acidic protein (non-histones) and also to Histone 1 (H1) (very lysine rich, F1). Two-hour DBD treatment resulted in partial degradation and reduced [3H]leucine incorporation into the same fractions of chromatin. However, 6 h after DBD treatment, the synthesis of the degraded chromatin proteins began and by 24 h was completed. During the same treatment period the composition of chromatin showed a remarkable alteration; 2 h after DBD treatment the amount of the nuclear residual acidic proteins relative to DNA decreased by approx. 50%, but returned to control value 24 h after drug treatment. This in conjunction with the data on [3H]leucine incorporation suggests that certain chromatin proteins are degraded and subsequently newly synthesised after DND treatment resulting in an exchange of chromatin components. The formation of a nucleohistone complex between H1 and DNA was inhibited by pretreatment of H1 and DBD, dianhydrodulcitol (DAD) and bischloroethylnitrosourea (BCNU).

Histone 1 phosphotransferase activities during the maturation of oocytes of Xenopus laevis

A characterization has been initiated of histone 1 (H1) phosphotransferases in oocytes, progesterone-treated oocytes, unfertilized eggs, and A-23187 ionophore-treated eggs of Xenopus laevis. With the exception of one activity that was detected only in the oocytes and which was characterized as free catalytic subunit of the cAMP-dependent protein kinase, the profile of H1 phosphotransferases activities was qualitatively identical in all four types of cells. The only cAMP-dependent protein kinase detected was the Type II isozyme. All four types of cells contain a H1 phosphotransferase which would appear to be very similar to the growth-associated H1 kinase identified in other eukaryotic cells on the basis of the criteria of (a) molecular size, (b) the phosphorylation of both serine and threonine residues in the C-terminal portion of H1, and (c) the phosphorylation of the synthetic peptides LysSerProValLys and LysThrProValLys. All four types of cells contain at least two other histone phosphotransferase activities, one of which exhibits a high degree of selectivity for H1.