Genomic analysis of the secretion stress response in the enzyme-producing cell factory Aspergillus niger

<p>Abstract</p> <p>Background</p> <p>Filamentous fungi such as <it>Aspergillus niger </it>have a high capacity secretory system and are therefore widely exploited for the industrial production of native and heterologous proteins. However, in most cases the yields of non-fungal proteins are significantly lower than those obtained for fungal proteins. One well-studied bottleneck appears to be the result of mis-folding of heterologous proteins in the ER during early stages of secretion, with related stress responses in the host, including the unfolded protein response (UPR). This study aims at uncovering transcriptional and translational responses occurring in <it>A. niger </it>exposed to secretion stress.</p> <p>Results</p> <p>A genome-wide transcriptional analysis of protein secretion-related stress responses was determined using Affymetrix DNA GeneChips and independent verification for selected genes. Endoplasmic reticulum (ER)-associated stress was induced either by chemical treatment of the wild-type cells with dithiothreitol (DTT) or tunicamycin, or by expressing a human protein, tissue plasminogen activator (t-PA). All of these treatments triggered the UPR, as shown by the expression levels of several well-known UPR target genes. The predicted proteins encoded by most of the up-regulated genes function as part of the secretory system including chaperones, foldases, glycosylation enzymes, vesicle transport proteins, and ER-associated degradation proteins. Several genes were down-regulated under stress conditions and these included several genes that encode secreted enzymes. Moreover, translational regulation under ER stress was investigated by polysomal fractionation. This analysis confirmed the post-transcriptional control of <it>hacA </it>expression and highlighted that differential translation also occurs during ER stress, in particular for some genes encoding secreted proteins or proteins involved in ribosomal biogenesis and assembly.</p> <p>Conclusion</p> <p>This is first genome-wide analysis of both transcriptional and translational events following protein secretion stress. Insight has been gained into the molecular basis of protein secretion and secretion-related stress in an effective protein-secreting fungus, and provides an opportunity to identify target genes for manipulation in strain improvement strategies.</p

Subdivision, interpolation and splines

Thesis (MSc)--University of Stellenbosch, 2000.ENGLISH ABSTRACT: In this thesis we study the underlying mathematical principles of stationary subdivision, which can be regarded as an iterative recursion scheme for the generation of smooth curves and surfaces in computer graphics. An important tool for our work is Fourier analysis, from which we state some standard results, and give the proof of one non-standard result. Next, since cardinal spline functions have strong links with subdivision, we devote a chapter to this subject, proving also that the cardinal B-splines are refinable, and that the corresponding Euler-Frobenius polynomial has a certain zero structure which has important implications in our eventual applications. The concepts of a stationary subdivision scheme and its convergence are then introduced, with as motivating example the de Rahm-Chaikin algorithm. Standard results on convergence and regularity for the case of positive masks are quoted and graphically illustrated. Next, we introduce the concept of interpolatory stationary subdivision, in which case the limit curve contains all the original control points. We prove a certain set of sufficient conditions on the mask for convergence, at the same time also proving the existence and other salient properties of the associated refinable function. Next, we show how the analysis of a certain Bezout identity leads to the characterisation of a class of symmetric masks which satisfy the abovementioned sufficient conditions. Finally, we show that specific special cases of the Bezout identity yield convergent interpolatory symmetric subdivision schemes which are identical to choosing the corresponding mask coefficients equal to certain point evaluations of, respectively, a fundamental Lagrange interpolation polynomial and a fundamental cardinal spline interpolant. The latter procedure, which is known as the Deslauriers-Dubuc subdivision scheme in the case of a polynomial interpolant, has received attention in recent work, and our approach provides a convergence result for such schemes in a more general framework. Throughout the thesis, numerical illustrations of our results are provided by means of graphs.AFRIKAANSE OPSOMMING: In hierdie tesis ondersoek ons die onderliggende wiskundige beginsels van stasionêre onderverdeling, wat beskou kan word as 'n iteratiewe rekursiewe skema vir die generering van gladde krommes en oppervlakke in rekenaargrafika. 'n Belangrike stuk gereedskap vir ons werk is Fourieranalise, waaruit ons sekere standaardresuJtate formuleer, en die bewys gee van een nie-standaard resultaat. Daarna, aangesien kardinale latfunksies sterk bande het met onderverdeling, wy ons 'n hoofstuk aan hierdie onderwerp, waarin ons ook bewys dat die kardinale B-Iatfunksies verfynbaar is, en dat die ooreenkomstige Euler-Frobenius polinoom 'n sekere nulpuntstruktuur het wat belangrike implikasies het in ons uiteindelike toepassings. Die konsepte van 'n stasionêre onderverdelingskema en die konvergensie daarvan word dan bekendgestel, met as motiverende voorbeeld die de Rahm-Chaikin algoritme. Standaardresultate oor konvergensie en regulariteit vir die geval van positiewe maskers word aangehaal en grafies geïllustreer. Vervolgens stelons die konsep van interpolerende stasionêre onderverdeling bekend, in welke geval die limietkromme al die oorspronklike kontrolepunte bevat. Ons bewys 'n sekere versameling van voldoende voorwaardes op die masker vir konvergensie, en bewys terselfdertyd die bestaan en ander toepaslike eienskappe van die ge-assosieerde verfynbare funksie. Daarna wys ons hoedat die analise van 'n sekere Bezout identiteit lei tot die karakterisering van 'n klas simmetriese maskers wat die bovermelde voldoende voorwaardes bevredig. Laastens wys ons dat spesifieke spesiale gevalle van die Bezout identiteit konvergente interpolerende simmetriese onderverdelingskemas lewer wat identies is daaraan om die ooreenkomstige maskerkoëffisientegelyk aan sekere puntevaluasies van, onderskeidelik, 'n fundamentele Lagrange interpolasiepolinoom en 'n kardinale latfunksie-interpolant te kies. Laasgenoemde prosedure, wat bekend staan as die Deslauriers-Dubuc onderverdelingskema in die geval van 'n polinoominterpolant, het aandag ontvang in onlangse werk, en ons benadering verskaf 'n konvergensieresultaat vir sulke skemas in 'n meer algemene raamwerk. Deurgaans in die tesis word numeriese illustrasies van ons resultate met behulp van grafieke verskaf

Is dat iets voor mij, juf? Leerlingen versterken in het keuzeproces van basis naar secundair

'Latijn is 't moeilijkste.' 'Ze hadden toch wel graag dat ik een "hoge" richting ging volgen.' 'TSO is iets voor jongens.' De overgang tussen de basisschool en het secundair onderwijs blijft een scharniermoment voor jongeren. Niet alleen moeten leerlingen dan een middelbare school kiezen, ze selecteren ook een pakket optie-uren die een sterke impact kunnen hebben op hun schoolloopbaan. Is dat iets voor mij, juf? onderzoekt die overstap, ontrafelt de initiatieven die basisscholen nemen om leerlingen en ouders erbij te begeleiden en gaat in op de ervaringen van leerlingen, leerkrachten en ouders. Welke rol spelen ze in het keuzeproces? Welke kansen en drempels ervaren ze? Welke (stereotiepe) ideeën leven er en hebben die een effect op het zelfbeeld van de leerling? Dit boek reikt concrete handvatten en aanbevelingen aan om de school- en studiekeuze zo vlot mogelijk te maken en het welbevinden van alle betrokkenen te verhogen. Zo werpt het boek een dam op tegen enkele pijnpunten, zoals het hardnekkige watervaldenken en de vaststelling dat leerlingen en ouders met de grootste noden zich te weinig ondersteund voelen