Cloning of Linker Histones - Essay Example

Cloning of Linker Histones Cloning of Linker Histones Question Histones are highly conserved basic proteins that associate with the DNA with a fixed stoichiometry to constitute the essential ‘nucleosome’ that packages the genomic DNA into respective compact structures. There exist five types of histones namely H2B, H2A, H1, H3, and H4 linker histone. They exist as a complex of (H32-H42) and two dimers of (H2A-H2B) ultimately forming an octamer within the nucleosome. The H1 variant associates externally with the nucleosome helping in further compaction of the chromatin structure. In total, the humans have 55 unique histone variants. Histones have a characteristic ‘histone fold domain’ consisting of structural motif known as ‘helix-turn-helix and that are three alpha helices that all connected by loops. Each of the histones fits perfectly with their counterpart to form heterodimer structures that assume the appearance of a hand-shake. The histone cell structures are buried inside the core structure of the nucleosome. The histones have conspicuous N-terminal tails protruding out of the compact structure. Often the terminal tails are subjected to several post-translational modification that include methylation, ubiquitination, acetylation, phosphorylation, and many more (Xie 2009). It is the combination of the marks they get through the modification processes, and that determine the factors that bind to the region of DNA and in the long run regulation the expression status of the given locus. The multiple loci occurring as distinct clusters on different chromosomes are where the histone genes typically transcribed. Histone proteins have their individual repertoire of variants distinct in the sequence of their amino acid mostly in the protruding N-terminal region. The expression of the variants, which is dependent on the type, can either be replication-dependent or replication-independent. As will be discussed in a dedicated section below, their main function is to mark specific regions of the DNA by replacing canonical histone from the nucleosomes present and the particular site. This stress on distinct regions in the genome has a noteworthy part in recruitment of diverse factors to that site occasioning differential treatment. It is this mechanism that lays the foundation for creation and development of an epigenetic ‘memory’. Question 2 The dyad axis of symmetry that is where the exiting and the entering DNA duplexes cross has been a long held outlook of the most likely location for the binding of the linker histone to the nucleosome core particles. The high concentrations of the backbone phosphates in this site generates a dense patch of negative charge which is essentially a good zone for the binding of the lysine-rich and arginine- protein through electrostatic interactions. Structural studies reveal that there exists a second potential binding site for DNA that is composed of a cluster of the highly conserved rudiment ally amino acids on the opposite face of the molecular. The observation was instrumental in explaining a few findings. Among them was the ability of the protein to form DNA molecules made up of tram-like structures and separated by a scaffold of linker histone molecules. The second finding was the preferential binding exhibited by the linker histones to crossovers of double-helical DNA and four-way artificial junctions over linear double-stranded DNA. The last finding was the ability it had to alter the path of the lender DNA so that it exists as a ‘stem’ from the nucleosome. Two full turns of DNA as a result of the linker histone binding lock on the surface of histone octamer which create a 168-base pair micrococci nuclease digestion intermediate and another 20 base pairs characteristically longer that the 146 base pairs of DNA typically protected by the nucleosome core. The two binding sites for the DNA in the globular domain are required for the formation of the digestion intermediate which suggests that it requires such an interaction of the proteins with two separate regions of the nucleosome DNA. The means that the linker histone generates asymmetric nuclease protection depends on several factors. Among them is where it binds nucleosomal DNA. The linker histone histones bind close to the aforementioned dyad axis of the nucleosome. The linker DNA may link one terminus of the nucleosomal dyad and the other terminus of the nucleosome. There are numerous studies that have come up with different observations on how they are located. Ura et al. (1995) postulate that the binding could result from off-dyad binding of the linker histone. The used photoactive derivation to identify where the linker histone may bind. The model they used came up with a proposal that the globular domain of the linker histone is bound inside the DNA gyres. More so, this binding was not at the center, but specifically on one side of the nucleosome particle they used. Question 3 The linker histone has many functions. The most outstanding is its participation in the condensation of the ‘beads-on-a-string’ fiber into higher structures and its ability to stabilize the nucleosomal particle structure. This notable correlation between the compacted structure of and the presence of linker histones on chromatin has led to the proposal that linker histones may function as indiscriminate repressors of transcription. The correlation of between the lender histones and the structure limits the access of components of the transcriptional machinery. The linker histones, more recently, have been said that they can regulated transcriptional at a finer level as they are capable of influencing the accessibility of specific transcription factors to their binding sites on particular promoters. Both the structural and the functional roles of the linker histone H1 remain a mystery yet they have received considerable attention for many years now. Earlier H1 concepts were generally those that focused on them as transcriptional inhibitors but have since been reconsidered as some experiments demonstrate minor effect of the H1 deletion in unicellular organisms. Recent research conducted on mice show that cells and tissue are able t tolerated incredibly low H1 content. The studies have been carried out through knockouts of selected H1 subtypes in the mammals. The H1, instead of making up the nucleosome “bead” sits on top of the structure and keeps the DNA that was apparently wrapped around the nucleosome in place (Cole 1987). Currently, it is still not clear whether the H1 promotes a solenoid DNA-like chromatin fiber (where the linker DNA is shortened) or just promotes an alteration in the angle of adjacent nucleosomes without affecting linker length. Still, the histone H1 protects about 15-30 base pairs of additional DNA. Question 4 The H1 family in animals consist of several H1 isoforms that can be expressed differently or overlapping tissues and developmental stages in the organisms. The reason behind this phenomenon remains unclear. Among the linker, histone variants are the isoform histone H5. This histone is only found in avian erythrocytes which have nuclei, unlike mammalian erythrocytes that have none. Another variant is the oocyte/zygotic H1M isoform that is also known as B4 or H1foo. This type of variant is found in a myriad of organisms that include frogs, sea urchins, mice, and humans replaced in the embryo by somatic isoforms H10, and H1-1 that resemble H5. In spite of these isoforms having more negative charges that somatic isoforms, the H1M binds with an even greater affinity to mitotic chromosomes in Xenopus egg extracts. H1 isoforms, at mitosis, undergo phosphorylation at multiple cyclin-dependent kinases 1 (CDK 1) consensus sites that introduce quite some negative charges in the process. In general, there are about eight different linker histone 1 (H1) variants. The variety found at a particular linker is dependent on the type of the cell of the linker, its stage of differentiation, and where it is in the cell cycle. Other examples of variants besides what has been mentioned above are H3 which is replaced by CENP-A, commonly known as centromere protein A, at the nucleosomes near centromeres. Then H2A replaced by the variant H2A.Z at gene enhancers and promoters. In general, all the “standard” histones are eventually replaced by variant as the sperm develop (Tachiwana 2010). References Cole, R. David. Microheterogeneity in H1 histones and its consequences. International journal of peptide and protein research 30.4, 1987 Tachiwana, Hiroaki, et al. Structural basis of instability of the nucleosome containing a testis-specific histone variant, human H3T. Proceedings of the National Academy of Sciences 107.23, 2010 Ura, Kiyoe, Jeffrey J. Hayes, and A. P. Wolffe. A positive role for nucleosome mobility in the transcriptional activity of chromatin templates: restriction by linker histones. The EMBO Journal 14.15, 1995 Xie, Wei, et al. Histone h3 lysine 56 acetylation is linked to the core transcriptional network in human embryonic stem cells. Molecular cell 33.4, 2009 Read More