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DEVELOPMENT OF MODELS FOR ELUCIDATING THE GENETIC BASIS AND PATHOGENESIS OF MITOCHONDRIAL DISEASES AND RELATED CONDITIONS
Full text linkThe yeast Saccharomyces cerevisiae is a cheap, time-saving, and versatile tool to study human
genetic diseases, since a great number of genes and biological processes are evolutionarily
conserved. Moreover, residues where mutations occur in human are often conserved in yeast. Not
least, yeast can grow either by mitochondrial-dependent respiration or by ethanol fermentation, thus
permitting to study mitochondrial diseases. We utilized this model organism to assess the functional
impact of mutations in STRADA, SGPL1, and COQ genes, which are linked to primary CoQ10
deficiency, and where our goal was also to explore and enhance our understanding of their
involvement in CoQ biosynthesis.
Loss of function in the STRADA gene leads to PMSE syndrome (MIM #611087), a rare
neurodevelopmental disorder characterized by drug-resistant epilepsy and severe developmental
delays. Treatment with sirolimus has been effective. We employed a yeast model to evaluate the
missense variants Ser264Arg in STRADA and its complete deletion, underscore the significance of
timely molecular diagnosis for these patients and highlight the utility of yeast as a straightforward yet
effective tool for validating the pathogenicity of missense variants.
The SGPL1 gene encodes an enzyme involved in sphingolipid metabolism. Yeast's similarity to the
human pathway makes it suitable for studying diseases like SPLIS (MIM # 617575), an autosomal
recessive syndrome characterized by steroid-resistant nephrotic syndrome and multisystemic
manifestations, resembling CoQ deficiency. Vitamin B6 shows promise in mitigating these diseases.
Our study focused on the Gly210Glu mutation, which is situated at the pyridoxal 5'-phosphate (PLP)
binding site (active form of vitamin B6) and revealed a potential role for the yeast gene GAD1.
Coenzyme Q (CoQ) biosynthesis involves multiple genes. We investigated PDSS1, PDSS2, COQ4,
and COQ5 in yeast.
In yeast, the COQ1 gene controls the length of the polyprenyl chain and the number of isoprene
units in CoQ synthesis. In contrast, humans and mice rely on PDSS1 and PDSS2 subunits to form
heterotetramers for CoQ biosynthesis. This study aimed to clarify the roles of polyprenyl diphosphate
genes and explore hybridization between human and murine sequences in determining isoprene
chain length in S.cerevisiae, where the orthologous protein typically forms homodimeric or
II
homotetrameric structures. We used a yeast model and revealed that both murine Pdss1 and Pdss2
effectively complemented the absence of the coq1 gene in S.cerevisiae. Thus, we also validated the
pathogenicity of 12 human missense mutations, 7 in PDSS1 (His78Asn, Glu123Gly, Val239Gly,
Gln245His, Gly296Arg, Asp308Glu, Ser370Arg) and 5 in PDSS2 (Val116Met, His162Arg,
Ser382Leu, Ala384Asp, Ile391Thr) genes. Additionally, we investigated mitochondrial import
capabilities of murine PDSS1 vs. human PDSS1, revealing the crucial role of the mPDSS1 Nterminal
in mitochondrial import in S.cerevisiae. Finally, we quantified CoQ levels in hybrid strains
using HPLC, confirming that PDSS1 subunit 1 primarily determines the number of isoprene units in
the Q-aromatic ring.
COQ4's role in CoQ synthesis remains unclear. We created a yeast model with coq4 deletion and
identified the importance of its metal-binding site, through the mutant Asp164Ala, and its catalytic
activity. We expressed wild-type and mutant COQ4 proteins in bacteria for future enzymatic assays.
COQ5 is crucial for CoQ synthesis and interacts with the CoQ-synthome. We confirmed genotypephenotype
correlations using yeast and studied severe and milder patient mutations, as Gly118Ser
and Arg123Trp.
Saccharomyces cerevisiae, a simple eukaryotic model, offers a well-annotated genome, rapid
generation times, and a versatile toolkit. These attributes position yeast as an optimal model for
understanding genes and mutations associated with human diseases
Dati preliminari sulle malte del teatro romano della città di Nora (Sardegna meridionale)
No full textIl rilievo integrale dell’area Tottubella. Sardegna nord-occidentale. Carta litologica (scala 1:25.000), Carta delle acclività (scala 1:50.000), Carta Morfologica (scala 1:50.000)
No full textA Multilevel bayesian model for contextual effect of material deprivation
No full textThe relationship between socioeconomic factors and health has been studied in many circumstances. Whether the association takes place at individual level only, or also at population level (contextual effect) is still unclear.We present a multilevel hierarchical Bayesian model to investigate the joint contribution of individual and population-based socioeconomic factors to mortality, using data from the census cohort of the general population of the city of Florence, Italy (Tuscany Longitudinal Study, 1991–1995). Evidence supporting a contextual effect of deprivation on mortality at the very fine level of aggregation is found. Inappropriate modelling of individual and aggregate variables could strongly bias effect estimates
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