Histone H2AX in the Study Term Paper

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These proteins include homologous members of yeast. The presences of these proteins suggest that E. histolytica is skilled to perform homologous recombination, which is the same as in other organisms. DNA damage was evaluated by TUNEL assay. In yeast and in human cells, histone H2AX becomes rapidly phosphorylated when DSBs are introduced into chromatin (Lavi et al.).

Studies show that histone as a protein plays a significant role in the transition between the expression of a fetal gene and that of the adult gene. The adult gene's metabolism becomes oxidative in order to adapt to air and to weight, as it generates methylated transmitters and creatine phosphate. The muscles get used to life on the ground as compared to the fetal life which takes place in an aquatic environment. Regulated proteins allow the muscles to respond in a more adequate manner to this environment.

Now, let us see how histone and histone-like proteins work in dinoflagellates, most of which are marine planktons that commonly live in fresh water and belongs to a large group of flagellate protists. When they reproduce, they generate red tide that contains toxins. According to BioMed Central, a significant finding of the study of histone and histone-llike proteins in dinoflagellates is the identification of two rare expressed sequence tags (ESTs) that encode a partial histone H2AX. The longest cDNA isolated form the library using PCR was predicted to encode a protein of 169 amino acids that share high sequence identity to eukaryotic histone H2AX. This clone putatively lacked only the start codon at the N-terminus. The divergent N-terminus of a. tamarense H2AX is somewhat longer than in other homologs but the remainder of the sequence is conserved. Several functional residues from the known crystal structure are also present in a. tamarense H2AX including the lysine at the trypsine cleavage site, the arginines in the loops that interact with the DNA ?-helix, and the lysine ubiquitination site (Luger: 1997). Here, H2AX functions in the recognition and repair of double-strand DNA breaks by non-homologous end-joining. Various studies have proven that H2AX can arise independently and in many times during eukaryotic evolution.

Crissman et al. made a cytochemical analyses of the nuclear histone composition of hepatic tissue from acid-exposed Brook trout. The liver sections were stained fro histones by the eosin-fast green method which has been used for detecting histone changes n other vertebrate species. Three spectrally distinct cell types were determined on the basis of differential affinities for eosin and fast green. They exposed the fish to low acidity or pH for four days and they found out that the exposure did not affect the relative proportions of the three cell types. It only indicates that the aspect of liver function is not influenced by short exposure to low pH (2001: 49-52).

Another study of the activity and cytological effects of histone-like proteins on the parasitic dinoflagellate Amyloodinium ocellatum were made by Noga et al. The authors isolated from the tissues of rainbow trout Oncorhynchus mykiss and hybrid striped bass some histone-like proteins (HLPs) that are broad spectrum that endogenously produced antibiotics. They found that the histone-like proteins, which have a high sequence homology to H2B, equally inhibited both young and mature trophonts of the important ectoparasite Amyloodinium ocellatum. They also found that the aside from direct killing of Amyloodinium trophonts, there was an evident that HLP-1 from both rainbow trout and hybrid striped bass caused severe developmental abnormalities, which include delayed development in both the parasitic trophont stage as well as reproductive tomont stage. Parasites of normal appearance would die later in development and this called the "delayed mortality" effect of HLP-1. They also have seen the same effect with calf histone H2B and unrelated peptide antibiotic magainin. The antibiotic activity in mucus vs. epidermis compartments of the skin of hybrid striped bass suggested that the majority of antibiotic, including HLP-1 activity, resided in the epidermis, although some activities were present in the mucus. With this, they concluded that normal, non-immune fish skin contains potent defenses against protozoan ectoparasites and that the effects of these defenses may extend beyond their transient interactions with the parasites, which has important implications for this host-parasite relationship (2002: 207-215). Further evidence that HLPs are important defensive molecules in fish is provided by the fact that the relatively mild extraction techniques which the authors used to obtain HLP-1 and HLP-2 from fish tissues are not strong enough to release histone bound to nuclei acid (Moehs et al. 1992).

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It was also found that the activity was just as strong when they extracted tissues in water (Robinette et al. 1998; E. Noga and Z. Fan, unpublished data). The most important thing in their study, was that they have found that HLP-1 and HLP-2 possess potent activity against important fish pathogens that include bacteria and water moulds (Robinette et al. 1998). As a result, HLPs and other endobiotics may prove to be a very rich source of anti-parasitic strategies in the future.

As a then PhD in Pharmacology student in MUSC, Severine Patat and her group made a research about shrimp innate immunity to identity antimicrobial peptides and proteins in shrimp hemolymp. Preliminary data showed that hemocytes or shrimp blood cells contain high levels of histone proteins. In their study, it was found that in various vertebrates, histones have been shown to have antimicrobial activity. Since hemocytes are immune effector cells in the shrimp, they hypothesized that hemocytes contained high levels of histones to fight microorganisms. Shrimp histones H2A, H2B/H4 and H1 fragment were purified and inhibited growth of the Gram positive bacteria M. luteus.

A research headed by Robinette et al. was made about "Antimicrobial activity in the skin of the channel catfish Ictalurus punctatus: characterization of broad-spectrum histone-like antimicrobial proteins." It is widely known that parasites are the major cause of morbidity and mortality in fish. (Schnick et al. 1997). Channel catfish skin contains three proteins with antimicrobial activity. This activity is acid -- and heat-stable, and highly basic in nature. They isolated three antibacterial proteins from acid extracts of channel catfish skin. Histone-like proteins (HLPs) from channel catfish which exhibit broad-expectrum activity against pathogenic bacteria and water moulds were isolated. The objective of the study was to determine if similar defences occurred in unrelated fish species and whether such antibiotics had activity against protozoan ectoparasites.

In their initial study, they obtained from the local farms some clinically normal rainbow trout and sunshine bass that range from thirty to forty five centimeters in length and about two to four kilograms in weight. These were processed on site. The fish were euthanized by a blow to the head and decapitation and then they were maintained on ice through the process. They scraped the epidermis from the dermis with a scalpel. The scrapings were placed into ice-cold 1% acetic acid. They also dissected from each fish the gill and spleen and applied the same process. Afterwards, they heated the suspension until it boiled, and then they allowed it to cool at room temperature. The material was then homogenized and they extracted the acid. laboratory results indicated that skin extracts from channel catfish had significant antibacterial activity. The process was made by cation exchange chromatography. They isolated the purity and apparent molecular mass of antibacterial proteins from RP-HPLC and were assessed using SDS-polyacrylamide gel electrophoresis. The molecular masses of the purified antibacterial proteins were determined by matrix-assisted laser desorption mass spectrometry and were measured at 15.5, 15.5 and 30 kD. The most abundant antibacterial protein was assayed for antibacterial activity using a spectrophotometric assay. It was confirmed that antimicrobial protein was accomplished using a modification of a gel overlay assay. Then they examined the antiparasitic activity with the use of standard in vitro assay. Since the previous data indicated that HLPs were highly basic molecules (Robinette et al. 1998), the authors produced a large amount of tissue extract in 1% acetic acid for further purification and then they exploited a method that was similar to that used in purifying histones (Moehs et al. 1992); Johns, 1977). Based on the gathered data on mass spectrometry, amino acid composition and amino acid sequence, the authors found that the sequence differed from the major antibacterial protein only by the substitution of lysine for glycine, with an approximately 82% homology with rainbow trout histone H2B. Because of their similarity to histones, the catfish antibacterial proteins that they isolated were designated histone-like proteins (HLP) 1,2, and 3 in decreasing order of predominance in RP-HPLC. HLP-1 is closely realated, if not identical, to histone H2B. The H2B-like protein was inhibitory to Aeromonas hydrohila and Saprolegnia spp., which are important bacterial and fungal pathogens of fish. Considering these findings, histones may be important defensive molecules in fish and functions in fish immunity. The authors also gathered other evidence that is suggestive of possible immunological roles for….....

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