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HUMAN GENETIC DISORDERS PART 2 – DIAGNOSIS AND TREATMENT
Get PDFThe first genetic test was run in 1978 by Kan and Dozy and independently by Orkin et al. The molecular testing was performed to detect the mutation c.20A>T in the β-globin DNA responsible for sickle cell anaemia. Kan and Dozy applied restriction analysis of the β-globin gene followed by hybridisation with a molecular probe complementary to the fragment of human β-globin sequence (RFLP method) [1]. Results enabled them the discrimination between mutant homozygotes (affected persons), normal homozygotes (healthy persons) and heterozygotes (mutation carriers). To detect the same mutation, Orkin et al. applied hybridisation with two variant oligonucleotide molecular probes to distinguish between mutant and normal alleles of the β-globin gene [2]. A milestone in development of molecular diagnostic techniques for detection of human mutations was the invention of polymerase chain reaction (PCR) made in 1983 by Kary Mullis. The PCR in molecular diagnostics is generally used to amplify DNA fragment before other molecular analyses are performed.
Genetic testing is divided in two categories: direct testing and indirect testing (gene tracking). Direct genetic testing involves testing for a certain genotype, i.e. identification of a specific pathogenic mutation within a gene, that is responsible for a certain condition. Indirect genetic testing also known as gene tracking tests the inheritance and segregation of genetic markers linked to a specific condition within a family
Identification of C-terminal hydrophobic residues important for dimerization and all known functions of ParB of Pseudomonas aeruginosa
Get PDFThe ParB protein of P. aeruginosa is important for growth, cell division, nucleoid segregation and different types of motility. To further understand its function we have demonstrated a vital role of the hydrophobic residues in the C-terminus of ParBP.a. By in silico modeling of the C-terminal domain (242-290 amino acids) the hydrophobic residues L282, V285 and I289 (but not L286) are engaged in leucine zipper-like structure formation, whereas the charged residues R290 and Q266 are implicated in forming a salt bridge involved in protein stabilization. Five parB mutant alleles were constructed and their functionality defined in vivo and in vitro. In agreement with model predictions the substitution of L286A had no effect on mutant protein activities. Two ParBs with single substitutions of L282A or V285A and deletions of two or seven C-terminal amino acids were impaired in both dimerization and DNA binding and were not able to silence genes adjacent to parS suggesting that dimerization through the C-terminus is a prerequisite for spreading on DNA. The defect in dimerization also correlated with loss of ability to interact with partner protein ParA. Reverse genetics demonstrated that a parB mutant producing ParB lacking the two C-terminal amino acids as well as mutants producing ParB with single substitution L282A or V285A had defects comparable to those of a parB null mutant. Thus so far all the properties of ParB seem to depend on dimerization
Nuclear Import and Export Signals of Human Cohesins SA1/STAG1 and SA2/STAG2 Expressed in Saccharomyces cerevisiae
Get PDFAbstract
Background: Human SA/STAG proteins, homologues of the yeast Irr1/Scc3 cohesin, are the least studied constituents of the
sister chromatid cohesion complex crucial for proper chromosome segregation. The two SA paralogues, SA1 and SA2, show
some specificity towards the chromosome region they stabilize, and SA2, but not SA1, has been shown to participate in
transcriptional regulation as well. The molecular basis of this functional divergence is unknown.
Methodology/Principal Findings: In silico analysis indicates numerous putative nuclear localization (NLS) and export (NES)
signals in the SA proteins, suggesting the possibility of their nucleocytoplasmic shuttling. We studied the functionality of
those putative signals by expressing fluorescently tagged SA1 and SA2 in the yeast Saccharomyces cerevisiae. Only the Nterminal
NLS turned out to be functional in SA1. In contrast, the SA2 protein has at least two functional NLS and also two
functional NES. Depending on the balance between these opposing signals, SA2 resides in the nucleus or is distributed
throughout the cell. Validation of the above conclusions in HeLa cells confirmed that the same N-terminal NLS of SA1 is
functional in those cells. In contrast, in SA2 the principal NLS functioning in HeLa cells is different from that identified in
yeast and is localized to the C-terminus.
Conclusions/Significance: This is the first demonstration of the possibility of non-nuclear localization of an SA protein. The
reported difference in the organization between the two SA homologues may also be relevant to their partially divergent
functions. The mechanisms determining subcellular localization of cohesins are only partially conserved between yeast and
human cells
Rsp5 Ubiquitin Ligase Is Required for Protein Trafficking in Saccharomyces cerevisiae COPI Mutants
Get PDFRetrograde trafficking from the Golgi to the endoplasmic reticulum (ER) depends on the formation of vesicles coated with the multiprotein complex COPI. In Saccharomyces cerevisiae ubiquitinated derivatives of several COPI subunits have been identified. The importance of this modification of COPI proteins is unknown. With the exception of the Sec27 protein (β’COP) neither the ubiquitin ligase responsible for ubiquitination of COPI subunits nor the importance of this modification are known. Here we find that the ubiquitin ligase mutation, rsp5-1, has a negative effect that is additive with ret1-1 and sec28Δ mutations, in genes encoding α- and ε-COP, respectively. The double ret1-1 rsp5-1 mutant is also more severely defective in the Golgi-to-ER trafficking compared to the single ret1-1, secreting more of the ER chaperone Kar2p, localizing Rer1p mostly to the vacuole, and increasing sensitivity to neomycin. Overexpression of ubiquitin in ret1-1 rsp5-1 mutant suppresses vacuolar accumulation of Rer1p. We found that the effect of rsp5 mutation on the Golgi-to-ER trafficking is similar to that of sla1Δ mutation in a gene encoding actin cytoskeleton proteins, an Rsp5p substrate. Additionally, Rsp5 and Sla1 proteins were found by co-immunoprecipitation in a complex containing COPI subunits. Together, our results show that Rsp5 ligase plays a role in regulating retrograde Golgi-to-ER trafficking
Impact of Yeast Glycosylation Pathway on Cell Integrity and Morphology, Glycosylation, Stefana Petrescu (Ed.), ISBN: 978-953-51-0771-2, InTech, Available from: http://www.intechopen.com/books/glycosylation/impact-of-yeast-glycosylation-pathway-on-cell-integrity-and-morphology
Get PDFProtein glycosylation is a multi step reaction, well conserved in the eukaryotic cells. In N-glycosylation reactions dolichyl phosphate (DolP) serves as a lipid acceptor of sugar residues forming DolPPGlcNAc2Man9Glc3. Dolichyl phosphate mannose (DolPMan) is also a substrate for protein O-glycosylation, where it serves as a donor of the first mannose to be attached to hydroxyl groups of serine or treonine. DolPMan is also involved in the synthesis of the sugar part of glycosylphosphatidyl inositol anchor in yeast and other eukaryotes. Its remnant structure is responsible for the attachment of a large group of glycoproteins to the glucan polymers of the cell wall . Thus, a functional link could be predicted between the dolichol biosynthetic (mevalonate) pathway and subsequent N-glycosylation and O-mannosylation, cell wall assembly and/or fungus–host interaction. Moreover, on the basis of the data presented in this chapter, it can be assumed that the glycosylation pathway in yeast and fungi offers many levels of regulation, which might influence the final quality and quantity of cell wall glycoproteins and consequently cell surface immunogenicity.
In this work we concentrate on early glycosylation defects, resulting from the impaired synthesis of dolichol (Dol) and dolichyl phosphate (DolP) or DolPP oligosaccharide (DolPPGlcNAc2Man9Glc3) assembly, and their effect on the cell integrity and morphology
Budowa, biogeneza i mechanizm działania kompleksu mitochondrialnej syntazy ATP
Get PDFMitochondria to organelle występujące u wszystkich organizmów eukariotycznych. Ich główną funkcją jest wytwarzanie energii w postaci ATP na drodze fosforylacji oksydacyjnej. Ostatni etap syntezy ATP katalizuje enzym wewnętrznej błony mitochondrialnej - syntaza ATP. Jest to kompleks złożony z co najmniej siedemnastu podjednostek (u drożdży, u kręgowców zidentyfikowano ich dotychczas szesnaście), tworzących część hydrofobową zagłębioną w błonie (nazwaną FO) i hydrofilową, skierowaną do macierzy mitochondrialnej (F1). Geny większości podjednostek znajdują się w genomie jądrowym, ale niektóre z nich, kodujące hydrofobowe podjednostki błonowe, u większości organizmów zachowane zostały w genomie mitochondrialnym. Biogeneza syntazy ATP jest procesem złożonym, wymaga bowiem udziału licznych białek niebędących podjednostkami enzymu, regulujących ekspresję genów syntazy oraz składanie podjednostek w dojrzały enzym. Niniejsze opracowanie stanowi podsumowanie aktualnego stanu wiedzy o budowie, biogenezie i mechanizmie działania kompleksu syntazy ATP
Coexistence of Gilbert syndrome with hereditary haemolytic anaemias.
Get PDFGenotyping of the UGT1A1 gene showed distinct distribution of the common A(TA)(n)TAA polymorphism relative to other European populations. Because of a greater risk of hyperbilirubinaemia due to hereditary haemolytic anaemia, the diagnosis of Gilbert syndrome in this group of patients is very important
Altered gene expression pattern in peripheral blood mononuclear cells in patients with acute myocardial infarction.
Get PDFIn the acute phase of STEMI, dozens of genes from several pathways linked with lipid/glucose metabolism, platelet function and atherosclerotic plaque stability show altered expression. Up-regulation of SOCS3 and FAM20 genes in the first days of myocardial infarction is observed in the vast majority of patients
The lichen biota of Antarctic Specially Protected Area No. 151, Lions Rump (King George Island)
Get PDFLions Rump lies on the southern coast of King George Bay on King George Island
in Antarctica. The part of the area, as an effect of Polish initiative, has been approved as protected area
(ASPA No. 151). The aim of this study was to document the occurrence and distribution of lichens
to provide the baseline data for long-term monitoring of the ongoing changes in this valuable area