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Fulltext Whole-Genome Analysis of Quorum-Sensing Burkholderia sp. Strain A9

Chan KG, Chen JW, Tee KK, Chang CY, Yin WF, Chan XY

Genome Announc, 2015;3(2).
PMID: 25745000 DOI: 10.1128/genomeA.00063-15

Burkholderia spp. rely on N-acyl homoserine lactone as quorum-sensing signal molecules which coordinate their phenotype at the population level. In this work, we present the whole genome of Burkholderia sp. strain A9, which enables the discovery of its N-acyl homoserine lactone synthase gene.

Matched MeSH terms: Ligases
Genome sequencing-assisted identification and the first functional validation of N-acyl-homoserine-lactone synthases from the Sphingomonadaceae family

Gan HM, Dailey LK, Halliday N, Williams P, Hudson AO, Savka MA

PeerJ, 2016;4:e2332.
PMID: 27635318 DOI: 10.7717/peerj.2332

Members of the genus Novosphingobium have been isolated from a variety of environmental niches. Although genomics analyses have suggested the presence of genes associated with quorum sensing signal production e.g., the N-acyl-homoserine lactone (AHL) synthase (luxI) homologs in various Novosphingobium species, to date, no luxI homologs have been experimentally validated.

Matched MeSH terms: Ligases
Fulltext Functional Characterisation of New Sesquiterpene Synthase from the Malaysian Herbal Plant, Polygonum Minus

Rusdi NA, Goh HH, Sabri S, Ramzi AB, Mohd Noor N, Baharum SN

Molecules, 2018 06 06;23(6).
PMID: 29882808 DOI: 10.3390/molecules23061370

Polygonum minus (syn. Persicaria minor) is a herbal plant that is well known for producing sesquiterpenes, which contribute to its flavour and fragrance. This study describes the cloning and functional characterisation of PmSTPS1 and PmSTPS2, two sesquiterpene synthase genes that were identified from P. minus transcriptome data mining. The full-length sequences of the PmSTPS1 and PmSTPS2 genes were expressed in the E. coli pQE-2 expression vector. The sizes of PmSTPS1 and PmSTPS2 were 1098 bp and 1967 bp, respectively, with open reading frames (ORF) of 1047 and 1695 bp and encoding polypeptides of 348 and 564 amino acids, respectively. The proteins consist of three conserved motifs, namely, Asp-rich substrate binding (DDxxD), metal binding residues (NSE/DTE), and cytoplasmic ER retention (RxR), as well as the terpene synthase family N-terminal domain and C-terminal metal-binding domain. From the in vitro enzyme assays, using the farnesyl pyrophosphate (FPP) substrate, the PmSTPS1 enzyme produced multiple acyclic sesquiterpenes of β-farnesene, α-farnesene, and farnesol, while the PmSTPS2 enzyme produced an additional nerolidol as a final product. The results confirmed the roles of PmSTPS1 and PmSTPS2 in the biosynthesis pathway of P. minus, to produce aromatic sesquiterpenes.

Matched MeSH terms: Ligases/genetics; Ligases/metabolism*; Ligases/chemistry
Genetic study of early-onset Parkinson's disease in the Malaysian population

Tay YW, Tan AH, Lim JL, Lohmann K, Ibrahim KA, Abdul Aziz Z, et al.

Parkinsonism Relat Disord, 2023 Jun;111:105399.
PMID: 37209484 DOI: 10.1016/j.parkreldis.2023.105399

BACKGROUND: About 5-10% of Parkinson's disease (PD) cases are early onset (EOPD), with several genes implicated, including GBA1, PRKN, PINK1, and SNCA. The spectrum and frequency of mutations vary across populations and globally diverse studies are crucial to comprehensively understand the genetic architecture of PD. The ancestral diversity of Southeast Asians offers opportunities to uncover a rich PD genetics landscape, and identify common regional mutations and new pathogenic variants.
OBJECTIVES: This study aimed to investigate the genetic architecture of EOPD in a multi-ethnic Malaysian cohort.
METHODS: 161 index patients with PD onset ≤50 years were recruited from multiple centers across Malaysia. A two-step approach to genetic testing was used, combining a next-generation sequencing-based PD gene panel and multiplex ligation-dependent probe amplification (MLPA).
RESULTS: Thirty-five patients (21.7%) carried pathogenic or likely pathogenic variants involving (in decreasing order of frequency): GBA1, PRKN, PINK1, DJ-1, LRRK2, and ATP13A2. Pathogenic/likely pathogenic variants in GBA1 were identified in thirteen patients (8.1%), and were also commonly found in PRKN and PINK1 (11/161 = 6.8% and 6/161 = 3.7%, respectively). The overall detection rate was even higher in those with familial history (48.5%) or age of diagnosis ≤40 years (34.8%). PRKN exon 7 deletion and the PINK1 p.Leu347Pro variant appear to be common among Malay patients. Many novel variants were found across the PD-related genes.
CONCLUSIONS: This study provides novel insights into the genetic architecture of EOPD in Southeast Asians, expands the genetic spectrum in PD-related genes, and highlights the importance of diversifying PD genetic research to include under-represented populations.

Matched MeSH terms: Ubiquitin-Protein Ligases/genetics
DNA splicing systems with at most two cutting sites of a non-palindromic restriction enzyme

Nurul Izzaty Ismail, Wan Heng Fong, Nor Haniza Sarmin

MATEMATIKA, 2019;35(2):129-137.
MyJurnal

The modelling of splicing systems is simulated by the process of cleaving and recombining DNA molecules with the presence of a ligase and restriction enzymes which are biologically called as endodeoxyribonucleases. The molecules resulting from DNA splicing systems are known as splicing languages. Palindrome is a sequence of strings that reads the same forward and backward. In this research, the splicing languages resulting from DNA splicing systems with one non-palindromic restriction enzyme are determined using the notation from Head splicing system. The generalisations of splicing languages for DNA splicing systems involving a cutting site and two non-overlapping cutting sites of one non-palindromic restriction enzyme are presented in the first and second theorems, respectively, which are proved using direct and induction methods. The result from the first theorem shows a trivial string which is the initial DNA molecule; while the second theorem determines a splicing language consisting of a set of resulting DNA molecules from the respective DNA splicing system.

Matched MeSH terms: Ligases
Fulltext Kesan sitotoksik sebatian difenilstanum (iv) alkilfenilditiokarbamat terhadap sel eritroleukemia, K562

Normah Awang, Siti Musslihah Shahidi, Asmah Hamid, Nurul Farahana Kamaludin

Jurnal Sains Kesihatan Malaysia, 2013;11(1):25-32.
MyJurnal

Kesan sitotoksik sebatian organostanum (IV) terhadap pelbagai sel kanser telah dikaji oleh para saintis di seluruh dunia.Dalam kajian ini,dua sebatian baru organostanum (IV) iaitu difenilstanum (IV) etilfenilditiokarbamat (DFEF) dan difenilstanum (IV) butilfenilditiokarbamat (DFBF) telah diuji kesan sitotoksiknya terhadap sel eritroleukemia, K562. Sel eritroleukemia, K562 merupakan sel sasaran manakala, sel hepar Chang dan sel fibroblas V79 pula digunakan untuk menilai kesan kedua-dua sebatian ini terhadap sel bukan kanser. Kesan sitotoksik sebatian DFEF dan DFBF diuji menggunakan ujian asai 3-(4,5-dimetiltiazol-2-il)-2, 5-difeniltetrazolium bromida (MTT) dengan masa pendedahan 24 jam, 48 jam dan 72 jam pada kepekatan sebatian yang berbeza. Pemerhatian terhadap perubahan morfologi juga dilakukan menggunakan nilai IC50 yang diperolehi pada masa pendedahan seperti ujian asai MTT. Ujian sitotoksisiti telah menunjukkan sebatian DFEF dan DFBF adalah sangat toksik terhadap sel K562 dengan nilai IC50 kurang daripada 10 μM untuk ketiga-tiga masa pendedahan.Indeks pemilihan juga membuktikan bahawa kedua-dua sebatian memberikan kesan sitotoksik secara memilih terhadap sel K562 pada masa 48 jam dan 72 jam, tetapi pada masa 24 jam, sebatian ini bertindak secara tidak memilih terhadap sel K562 dan sel bukan kanser. Perubahan morfologi yang diperhatikan adalah menyerupai ciri-ciri apoptosis seperti pengecutan sel dan pembentukan jasad apoptotik dan juga nekrosis seperti sel lisis. Kesimpulannya, sebatian difenilstanum (IV) alkilfenilditiokarbamat berpotensi untuk dibangunkan sebagai agen antileukemia tetapi mekanisma khusus tindakan sebatian ini terhadap sel K562 perlu dikaji pada masa akan datang untuk menjelaskan potensi sebatian ini sebagai dadah antikanser yang baru.

Matched MeSH terms: Ubiquitin-Protein Ligases
Fulltext Generalisations of DNA splicing systems with one palindromic restriction enzyme

Wan, Heng Fong, Nurul Izzaty Ismail

MATEMATIKA, 2018;34(1):59-71.
MyJurnal

In DNA splicing system, the potential effect of sets of restriction enzymes and
a ligase that allow DNA molecules to be cleaved and re-associated to produce further
molecules is modelled mathematically. This modelling is done in the framework of formal
language theory, in which the nitrogen bases, nucleotides and restriction sites are modelled
as alphabets, strings and rules respectively. The molecules resulting from a splicing system
is depicted as the splicing language. In this research, the splicing language resulting from
DNA splicing systems with one palindromic restriction enzyme for one and two (nonoverlapping)
cutting sites are generalised as regular expressions.

Matched MeSH terms: Ligases
Structural insights on Mycobacterium tuberculosis thiazole synthase--a molecular dynamics/docking approach

Rohini K, Srikumar PS

Appl Biochem Biotechnol, 2013 Mar;169(6):1790-8.
PMID: 23340867 DOI: 10.1007/s12010-013-0110-9

Tuberculosis (TB), an epidemic disease, affects the world with death rate of two million people every year. The bacterium Mycobacterium tuberculosis was found to be a more potent and disease-prolonged bacterium among the world due to multi-drug resistance. Emergence of new drug targets is needed to overcome the bacterial resistance that leads to control epidemic tuberculosis. The pathway thiamine biosynthesis was targeting M. tuberculosis due to its role in intracellular growth of the bacterium. The screening of enzymes involved in thiamin biosynthesis showed novel target thiazole synthase (ThiG) involved in catalysis of rearrangement of 1-deoxy-D-xylulose 5-phosphate (DXP) to produce the thiazole phosphate moiety of thiamine. We carried out homology modeling for ThiG to understand the structure-function relationship, and the model was refined with MD simulations. The results showed that the model predicted with (α + β)8-fold of synthase family proteins. Molecular docking of ThiG model with substrate DXP showed binding mode and key residues ARG46, ASN69, THR41, and LYS96 involved in the catalysis. First-line anti-tuberculosis drugs were docked with ThiG to identify the inhibition. The report showed the anti-tuberculosis drugs interact well with ThiG which may lead to block thiamin biosynthesis pathway.

Matched MeSH terms: Ligases/metabolism*; Ligases/chemistry*
ROCK2 inhibition: A futuristic approach for the management of Alzheimer's disease

Mani S, Jindal D, Chopra H, Jha SK, Singh SK, Ashraf GM, et al.

Neurosci Biobehav Rev, 2022 11;142:104871.
PMID: 36122738 DOI: 10.1016/j.neubiorev.2022.104871

Neurons depend on mitochondrial functions for membrane excitability, neurotransmission, and plasticity. Mitochondrial dynamics are important for neural cell maintenance. To maintain mitochondrial homeostasis, lysosomes remove dysfunctional mitochondria through mitophagy. Mitophagy promotes mitochondrial turnover and prevents the accumulation of dysfunctional mitochondria. In many neurodegenerative diseases (NDDs), including Alzheimer's disease (AD), mitophagy is disrupted in neurons. Mitophagy is regulated by several proteins; recently, Rho-associated coiled-coil containing protein kinase 2 (ROCK2) has been suggested to negatively regulate the Parkin-dependent mitophagy pathway. Thus, ROCK2 inhibition may be a promising therapy for NDDs. This review summarizes the mitophagy pathway, the role of ROCK2 in Parkin-dependent mitophagy regulation, and mitophagy impairment in the pathology of AD. We further discuss different ROCK inhibitors (synthetic drugs, natural compounds, and gene therapy-based approaches) and examine their effects on triggering neuronal growth and neuroprotection in AD and other NDDs. This comprehensive overview of the role of ROCK in mitophagy inhibition provides a possible explanation for the significance of ROCK inhibitors in the therapeutic management of AD and other NDDs.

Matched MeSH terms: Ubiquitin-Protein Ligases/genetics; Ubiquitin-Protein Ligases/metabolism
MicroRNA regulation of CTP synthase and cytoophidium in Drosophila melanogaster

Dzaki N, Woo WK, Thangadurai S, Azzam G

Exp Cell Res, 2019 12 15;385(2):111688.
PMID: 31678212 DOI: 10.1016/j.yexcr.2019.111688

CTPsyn is a crucial metabolic enzyme which synthesizes CTP nucleotides. It has the extraordinary ability to compartmentalize into filaments termed cytoophidia. Though the structure is evolutionarily conserved across kingdoms, the mechanisms behind their formation remain unknown. MicroRNAs (miRNAs) are short single-stranded RNA capable of directing mRNA silencing and degradation. D. melanogaster has a high total gene count to miRNA gene number ratio, alluding to the possibility that CTPsyn too may come under their regulation. A thorough miRNA overexpression involving 123 miRNAs was conducted, followed by CTPsyn-specific staining upon cytoophidia-rich egg chambers. This revealed a small group of candidates which confer either a lengthening or truncating effect on cytoophidia, suggesting they may play a role in regulating CTPsyn. MiR-975 and miR-1014 are both cytoophidia-elongating, whereas miR-190 and miR-932 are cytoophidia-shortening. Though target prediction shows that miR-975 and miR-932 do indeed have binding sites on CTPsyn mRNA, in vitro assays instead revealed a low probability of this being true, instead indicating that the effects asserted by overexpressed miRNAs indirectly reach CTPsyn and its cytoophidia through the actions of middling elements. In silico target prediction and qPCR quantification indicated that, at least for miR-932 and miR-1014, these undetermined elements may be players in fat metabolism. This is the first study to thoroughly investigate miRNAs in connection to CTPsyn expression and activity in any species. The findings presented could serve as a basis for further queries into not only the fundamental aspects of the enzyme's regulation, but may uncover new facets of closely related pathways as well.

Matched MeSH terms: Carbon-Nitrogen Ligases/genetics; Carbon-Nitrogen Ligases/metabolism*
Fulltext Unravelling the genetic links between Parkinson's disease and lung cancer

Leong YQ, Koh RY, Chye SM, Ng KY

Biol Chem, 2023 May 25;404(6):551-567.
PMID: 36634094 DOI: 10.1515/hsz-2022-0228

Increase evidence from epidemiological studies have shown an inverse association between Parkinson's disease (PD) and lung cancer. PD and lung cancer are both geriatric diseases, where these two diseases are sharing some common genetic determinants. Several PD-associated genes including alpha synuclein (SNCA), PTEN-induced kinase 1 (PINK1), parkin, parkinsonism associated deglycase (DJ-1), leucine-rich repeat kinase 2 (LRRK2), F-box protein 7 (FBXO7) and ubiquitin C-terminal hydrolase L1 (UCHL1) were reported to have altered expressions in lung cancer patients. This indicates that certain PD-associated genes might be important in conferring anticancer effects. This review aims to depict the physiological functions of these genes, and discuss the putative roles of these PD-associated genes in lung cancer. The understanding of the roles of these genes in the lung cancer progression might be important in the identification of new treatment targets for lung cancer. Gene therapy that aims to alter the expressions of these genes could be developed for future anticancer therapy. As a result, studying the roles of these genes in lung cancer may also help to understand their involvements as well as their roles in the pathogenesis of PD.

Matched MeSH terms: Ubiquitin-Protein Ligases/genetics; Ubiquitin-Protein Ligases/metabolism
The chloroplast-associated protein degradation pathway controls chromoplast development and fruit ripening in tomato

Ling Q, Sadali NM, Soufi Z, Zhou Y, Huang B, Zeng Y, et al.

Nat Plants, 2021 05;7(5):655-666.
PMID: 34007040 DOI: 10.1038/s41477-021-00916-y

The maturation of green fleshy fruit to become colourful and flavoursome is an important strategy for plant reproduction and dispersal. In tomato (Solanum lycopersicum) and many other species, fruit ripening is intimately linked to the biogenesis of chromoplasts, the plastids that are abundant in ripe fruit and specialized for the accumulation of carotenoid pigments. Chromoplasts develop from pre-existing chloroplasts in the fruit, but the mechanisms underlying this transition are poorly understood. Here, we reveal a role for the chloroplast-associated protein degradation (CHLORAD) proteolytic pathway in chromoplast differentiation. Knockdown of the plastid ubiquitin E3 ligase SP1, or its homologue SPL2, delays tomato fruit ripening, whereas overexpression of SP1 accelerates ripening, as judged by colour changes. We demonstrate that SP1 triggers broader effects on fruit ripening, including fruit softening, and gene expression and metabolism changes, by promoting the chloroplast-to-chromoplast transition. Moreover, we show that tomato SP1 and SPL2 regulate leaf senescence, revealing conserved functions of CHLORAD in plants. We conclude that SP1 homologues control plastid transitions during fruit ripening and leaf senescence by enabling reconfiguration of the plastid protein import machinery to effect proteome reorganization. The work highlights the critical role of chromoplasts in fruit ripening, and provides a theoretical basis for engineering crop improvements.

Matched MeSH terms: Ubiquitin-Protein Ligases/metabolism; Ubiquitin-Protein Ligases/physiology
CTP synthase knockdown during early development distorts the nascent vertebral column and causes fluid retention in multiple tissues in zebrafish

Dzaki N, Wahab W, Azlan A, Azzam G

Biochem Biophys Res Commun, 2018 10 20;505(1):106-112.
PMID: 30241946 DOI: 10.1016/j.bbrc.2018.09.074

CTP Synthase (CTPS) is a metabolic enzyme that is recognized as a catalyst for nucleotide, phospholipid and sialoglycoprotein production. Though the structural characteristics and regulatory mechanisms of CTPS are well-understood, little is known regarding the extent of its involvement during the early developmental stages of vertebrates. Zebrafish carries two CTPS genes, annotated as ctps1a and ctps1b. Phylogenetic analyses show that both genes had diverged from homologues in the ancestral Actinopterygii, Oreochromis niloticus. Conservation of common CTPS-catalytic regions further establishes that both proteins are likely to be functionally similar to hsaCTPS. Here, we show that ctps1a is more critical throughout the initial period of embryonic development than ctps1b. The effects of concurrent partial knockdown are dependent on ctps1a vs ctps1b dosage ratios. When these are equally attenuated, abnormal phenotypes acquired prior to the pharyngula period disappear in hatchlings (48hpf); however, if either gene is more attenuated than the other, these only become more pronounced in advanced stages. Generally, disruption to normal ctps1a or ctps1b expression levels by morpholinos culminates in the distortion of the early spinal column as well as multiple-tissue oedema. Other effects include slower growth rates, increased mortality rates and impaired structural formation of the young fish's extremities. Embryos grown in DON, a glutamine-analogue drug and CTPS antagonist, also exhibit similar characteristics, thus strengthening the validity of the morpholino-induced phenotypes observed. Together, our results demonstrate the importance of CTPS for the development of zebrafish embryos, as well as a disparity in activity and overall importance amongst isozymes.

Matched MeSH terms: Carbon-Nitrogen Ligases/classification; Carbon-Nitrogen Ligases/genetics*; Carbon-Nitrogen Ligases/metabolism
Analysis of core protein clusters identifies candidate variable sites conferring metronidazole resistance in Helicobacter pylori

Chua EG, Debowski AW, Webberley KM, Peters F, Lamichhane B, Loke MF, et al.

Gastroenterol Rep (Oxf), 2019 Feb;7(1):42-49.
PMID: 30792865 DOI: 10.1093/gastro/goy048

Background: Metronidazole is one of the first-line drugs of choice in the standard triple therapy used to eradicate Helicobacter pylori infection. Hence, the global emergence of metronidazole resistance in Hp poses a major challenge to health professionals. Inactivation of RdxA is known to be a major mechanism of conferring metronidazole resistance in H. pylori. However, metronidazole resistance can also arise in H. pylori strains expressing functional RdxA protein, suggesting that there are other mechanisms that may confer resistance to this drug.
Methods: We performed whole-genome sequencing on 121 H. pylori clinical strains, among which 73 were metronidazole-resistant. Sequence-alignment analysis of core protein clusters derived from clinical strains containing full-length RdxA was performed. Variable sites in each alignment were statistically compared between the resistant and susceptible groups to determine candidate genes along with their respective amino-acid changes that may account for the development of metronidazole resistance in H. pylori.
Results: Resistance due to RdxA truncation was identified in 34% of metronidazole-resistant strains. Analysis of core protein clusters derived from the remaining 48 metronidazole-resistant strains and 48 metronidazole-susceptible identified four variable sites significantly associated with metronidazole resistance. These sites included R16H/C in RdxA, D85N in the inner-membrane protein RclC (HP0565), V265I in a biotin carboxylase protein (HP0370) and A51V/T in a putative threonylcarbamoyl-AMP synthase (HP0918).
Conclusions: Our approach identified new potential mechanisms for metronidazole resistance in H. pylori that merit further investigation.

Matched MeSH terms: Carbon-Nitrogen Ligases
Agrobacterium tumefaciens mediated transformation of the proteolysis 6 (prt6) gene into cotyledons of tomato cv. Micro tom

Intan Elya Suka, Nur Farhana Roslan, Zamri Zainal, Nurulhikma Md Isa, Bee LC

Sains Malaysiana, 2018;47:1465-1471.

Gen Proteolisis 6 (PRT6) merupakan gen yang memainkan peranan penting dalam tapak jalan N-end rule dan berfungsi
sebagai enzim E3 ligase. PRT6 berperanan dalam pengenalan protein sasaran bagi proses degradasi. Objektif utama kajian
ini adalah untuk mentransformasi konstruk RNAi PRT6 ke dalam tomato berperantarakan Agrobacterium tumefaciens.
Ini bertujuan untuk memahami peranan tapak jalan N-end rule semasa proses pemasakan buah. Beberapa faktor yang
memberi kesan kepada transformasi seperti masa ko-penanaman dan juga kepekatan antibiotik yang digunakan telah
dioptimumkan. Keputusan kajian menunjukkan pengeraman kotiledon selama 48 jam pada medium ko-penanaman dapat
meningkatkan penghasilan kalus sebanyak 61% manakala penggunaan 500 mg/L antibiotik karbenisilin dalam medium
regenerasi pucuk dapat mengurangkan kontaminasi A. tumefaciens sehingga 5.2%. Selain itu, strain A. tumefaciens
C58 merupakan strain A. tumefaciens yang paling sesuai digunakan sebagai perantara dalam kajian ini. Tindak balas
berantai polimerase (PCR) telah dijalankan pada pucuk yang terhasil untuk mengesahkan integrasi fragmen PRT6 ke dalam
genom tomato. Berdasarkan analisis PCR, kesemua tujuh pucuk putatif transgenik adalah merupakan transforman positif.

Matched MeSH terms: Ubiquitin-Protein Ligases
Fulltext Effects of Nephrolithiasis on Serum DNase (Deoxyribonuclease I and II) Activity and E3 SUMO-Protein Ligase NSE2 (NSMCE2) in Malaysian Individuals

Yusof F, Mehde AA, Mehdi WA, Raus RA, Ghazali H, Rahman AA

Biomed Environ Sci, 2015 Sep;28(9):660-5.
PMID: 26464253 DOI: 10.3967/bes2015.092

OBJECTIVE: Nephrolithiasis is one of the most common disorders of the urinary tract. The aim of this study was to examine a possible relationship between DNase I/II activity and E3 SUMO-protein ligase NSE2 in the sera of nephrolithiasis patients to evaluate the possibility of a new biomarker for evaluating kidney damage.
METHODS: Sixty nephrolithiasis patients and 50 control patients were enrolled in a case-control study. Their blood urea, creatinine, protein levels and DNase I/II activity levels were measured by spectrometry. Serum NSMCE2 levels were measured by ELISA. Blood was collected from patients of the government health clinics in Kuantan-Pahang and fulfilled the inclusion criteria.
RESULTS: The result indicated that mean levels of sera NSMCE2 have a significantly increase (P<0.01) in patients compared to control group. Compared with control subjects, activities and specific activities of serum DNase I and II were significantly elevated in nephrolithiasis patients (P$lt;0.01).
CONCLUSION: This study suggests that an increase in serum concentrations of DNase I/II and E3 SUMO-protein ligase NSE2 level can be used as indicators for the diagnosis of kidney injury in patients with nephrolithiasis.

Matched MeSH terms: Ligases/blood*
Fulltext Ubiquitylation of the ER-Shaping Protein Lunapark via the CRL3KLHL12 Ubiquitin Ligase Complex

Yuniati L, Lauriola A, Gerritsen M, Abreu S, Ni E, Tesoriero C, et al.

Cell Rep, 2020 05 19;31(7):107664.
PMID: 32433973 DOI: 10.1016/j.celrep.2020.107664

Cullin-RING ligases (CRLs) control key cellular processes by promoting ubiquitylation of a multitude of soluble cytosolic and nuclear proteins. Subsets of CRL complexes are recruited and activated locally at cellular membranes; however, few CRL functions and substrates at these distinct cellular compartments are known. Here, we use a proteomic screen to identify proteins that are ubiquitylated at cellular membranes and found that Lunapark, an endoplasmic reticulum (ER)-shaping protein localized to ER three-way junctions, is ubiquitylated by the CRL3KLHL12 ubiquitin ligase. We demonstrate that Lunapark interacts with mechanistic target of rapamycin complex-1 (mTORC1), a central cellular regulator that coordinates growth and metabolism with environmental conditions. We show that mTORC1 binds Lunapark specifically at three-way junctions, and lysosomes, where mTORC1 is activated, make contact with three-way junctions where Lunapark resides. Inhibition of Lunapark ubiquitylation results in neurodevelopmental defects indicating that KLHL12-dependent ubiquitylation of Lunapark is required for normal growth and development.

Matched MeSH terms: Ubiquitin-Protein Ligases/metabolism*
Fulltext Ectopic miR-975 induces CTP synthase directed cell proliferation and differentiation in Drosophila melanogaster

Woo WK, Dzaki N, Thangadurai S, Azzam G

Sci Rep, 2019 Apr 15;9(1):6096.
PMID: 30988367 DOI: 10.1038/s41598-019-42369-6

CTP synthase (CTPSyn) is an essential metabolic enzyme, synthesizing precursors required for nucleotides and phospholipids production. Previous studies have also shown that CTPSyn is elevated in various cancers. In many organisms, CTPSyn compartmentalizes into filaments called cytoophidia. In Drosophila melanogaster, only its isoform C (CTPSynIsoC) forms cytoophidia. In the fruit fly's testis, cytoophidia are normally seen in the transit amplification regions close to its apical tip, where the stem-cell niche is located, and development is at its most rapid. Here, we report that CTPSynIsoC overexpression causes the lengthening of cytoophidia throughout the entirety of the testicular body. A bulging apical tip is found in approximately 34% of males overexpressing CTPSynIsoC. Immunostaining shows that this bulged phenotype is most likely due to increased numbers of both germline cells and spermatocytes. Through a microRNA (miRNA) overexpression screen, we found that ectopic miR-975 concurrently increases both the expression levels of CTPSyn and the length of its cytoophidia. The bulging testes phenotype was also recovered at a penetration of approximately 20%. However, qPCR assays reveal that CTPSynIsoC and miR-975 overexpression each provokes a differential response in expression of a number of cancer-related genes, indicating that the shared CTPSyn upregulation seen in either case is likely the cause of observed testicular overgrowth. This study presents the first instance of consequences of miRNA-asserted regulation upon CTPSyn in D. melanogaster, and further reaffirms the enzyme's close ties to germline cells overgrowth.

Matched MeSH terms: Carbon-Nitrogen Ligases/metabolism*
Quercetin glycosides induced neuroprotection by changes in the gene expression in a cellular model of Parkinson's disease

Magalingam KB, Radhakrishnan A, Ramdas P, Haleagrahara N

J Mol Neurosci, 2015 Mar;55(3):609-17.
PMID: 25129099 DOI: 10.1007/s12031-014-0400-x

Quercetin glycosides, rutin and isoquercitrin, are potent antioxidants that have been found to possess neuroprotective effect in diseases like Parkinson's and Alzheimer's disease. In the present study, we have examined the gene expression changes with rutin and isoquercitrin pretreatment on 6-hydroxydopamine (6-OHDA)-treated toxicity in rat pheochromocytoma (PC12) cells. PC12 cells were pretreated with rutin or isoquercitrin and subsequently exposed to 6-OHDA. Rutin-pretreated PC12 attenuated the Park2, Park5, Park7, Casp3, and Casp7 genes which were expressed significantly in the 6-OHDA-treated PC12 cells. Rutin upregulated the TH gene which is important in dopamine biosynthesis, but isoquercitrin pretreatment did not affect the expression of this gene. Both rutin and isoquercitrin pretreatments upregulated the ion transport and antiapoptotic genes (NSF and Opa1). The qPCR array data were further validated by qRT-PCR using four primers, Park5, Park7, Casp3, and TH. This finding suggests that changes in the expression levels of transcripts encoded by genes that participate in ubiquitin pathway and dopamine biosynthesis may be involved in Parkinson's disease.

Matched MeSH terms: Ubiquitin-Protein Ligases/genetics; Ubiquitin-Protein Ligases/metabolism*
Identification of germline alterations in breast cancer predisposition genes among Malaysian breast cancer patients using panel testing

Ng PS, Wen WX, Fadlullah MZ, Yoon SY, Lee SY, Thong MK, et al.

Clin Genet, 2016 10;90(4):315-23.
PMID: 26757417 DOI: 10.1111/cge.12735

Although an association between protein-truncating variants and breast cancer risk has been established for 11 genes, only alterations in BRCA1, BRCA2, TP53 and PALB2 have been reported in Asian populations. Given that the age of onset of breast cancer is lower in Asians, it is estimated that inherited predisposition to breast cancer may be more significant. To determine the potential utility of panel testing, we investigated the prevalence of germline alterations in 11 established and 4 likely breast cancer genes in a cross-sectional hospital-based cohort of 108 moderate to high-risk breast cancer patients using targeted next generation sequencing. Twenty patients (19%) were identified to carry deleterious mutations, of whom 13 (12%) were in the BRCA1 or BRCA2, 6 (6%) were in five other known breast cancer predisposition genes and 1 patient had a mutation in both BRCA2 and BARD1. Our study shows that BRCA1 and BRCA2 account for the majority of genetic predisposition to breast cancer in our cohort of Asian women. Although mutations in other known breast cancer genes are found, the functional significance and breast cancer risk have not yet been determined, thus limiting the clinical utility of panel testing in Asian populations.

Matched MeSH terms: Ubiquitin-Protein Ligases/genetics; Ubiquitin-Protein Ligases/chemistry

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