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[{"key": "dc.contributor.advisor", "value": "Yl\u00e4nne, Jari", "language": null, "element": "contributor", "qualifier": "advisor", "schema": "dc"}, {"key": "dc.contributor.advisor", "value": "Lappalainen, Pekka", "language": null, "element": "contributor", "qualifier": "advisor", "schema": "dc"}, {"key": "dc.contributor.advisor", "value": "K\u00f6rber, Sarah", "language": null, "element": "contributor", "qualifier": "advisor", "schema": "dc"}, {"key": "dc.contributor.author", "value": "Vahvaselk\u00e4, Lotta", "language": null, "element": "contributor", "qualifier": "author", "schema": "dc"}, {"key": "dc.date.accessioned", "value": "2025-04-16T11:18:47Z", "language": null, "element": "date", "qualifier": "accessioned", "schema": "dc"}, {"key": "dc.date.available", "value": "2025-04-16T11:18:47Z", "language": null, "element": "date", "qualifier": "available", "schema": "dc"}, {"key": "dc.date.issued", "value": "2025", "language": null, "element": "date", "qualifier": "issued", "schema": "dc"}, {"key": "dc.identifier.uri", "value": "https://jyx.jyu.fi/handle/123456789/101481", "language": null, "element": "identifier", "qualifier": "uri", "schema": "dc"}, {"key": "dc.description.abstract", "value": "ZNF185 on yksi huonoimmin karakterisoiduista LIM-domeeni-proteiineista ja siit\u00e4 l\u00f6ytyy ainoastaam 37 artikkelia, joista vain muutama keskittyy ZNF185:n solutoimintoihin. LIM-domeeni-proteiinit osallistuvat yleens\u00e4 proteiinien v\u00e4lisiin vuorovaikutuksiin ja monet niist\u00e4 on yhdistetty erityisesti solun tukirankana toimivaan aktiiniin. T\u00e4ss\u00e4 pro gradu -tutkielmassa selvitettiin ZNF185:n roolia soluissa inaktivoimalla ZNF185-geeni CRISPR/Cas9- menetelm\u00e4n avulla. Geenin onnistunut inaktivointi varmennettiin sek\u00e4 genomiett\u00e4 proteiinitasolla ja yhteens\u00e4 viidess\u00e4 kloonissa ZNF185:n inaktivaatio oli t\u00e4ysin onnistunut. N\u00e4iden viiden kloonin lis\u00e4ksi tunnistettiin joitain osittaisia poistoklooneja, jotka ilmensiv\u00e4t vain pienempi\u00e4 ZNF185:n isoformeja. Perustuen U-2 OS-villityypiss\u00e4 tehtyihin ZNF185-vasta-ainev\u00e4rj\u00e4yksiin ja eksogeenisesti ilmennettyyn ZNF185:een, sen solunsis\u00e4inen sijainti pystyttiin m\u00e4\u00e4ritt\u00e4m\u00e4\u00e4n dorsaalisiin stressis\u00e4ikeisiin sek\u00e4 my\u00f6s heikosti ventraalisiin stressis\u00e4ikeisiin. ZNF185-poistogeenisten kloonien vasta-ainev\u00e4rj\u00e4yksiss\u00e4 dorsaaliset stressis\u00e4ikeet olivat himmeit\u00e4 tai puuttuivat kokonaan, mik\u00e4 sopii hyvin yhteen ZNF185:n solunsis\u00e4isen sijainnin kanssa. Lis\u00e4ksi vasta-ainev\u00e4rj\u00e4ykset paljastivat, ett\u00e4 solujen fokaaliadheesiot olivat morfologialtaan pidemm\u00e4t ja kooltaan suuremmat verrattuna villityypin solujen adheesioihin. ZNF185 inaktivaation seurauksena solut muodostivat my\u00f6s ep\u00e4normaalin pitk\u00e4ik\u00e4isi\u00e4 h\u00e4nt\u00e4rakenteita. Lis\u00e4ksi solujen etureuna oli ep\u00e4s\u00e4\u00e4nn\u00f6llisen muotoinen. Poistogeenisten solukloonien suuntavaistoa sek\u00e4 liikkumisnopeutta analysointiin satunnaisessa solun liikkumiskokeessa, ja tulosten mukaan kloonit olivat huomattavasti hitaampia kuin villityypin solut, mik\u00e4 voisi johtua suuremmista fokaaliadheesioista. ZNF185:n poistamisella ei kuitenkaan ollut suurta vaikutusta solujen suuntavaistoon. My\u00f6s RhoA-aktiivisuus mitattiin poistogeenisist\u00e4 klooneista ja villityypin soluista aikaisemman tutkimuksen varmentamiseksi, mutta n\u00e4ytteiden v\u00e4lill\u00e4 ei ollut suuria eroavaisuuksia. Kaikki ker\u00e4tty data viittaisi siihen, ett\u00e4 ZNF185:n poistamisen seurauksena solukuori on heikko tai h\u00e4iriintynyt rakenteeltaan, ja poistogeeniset kloonit yritt\u00e4v\u00e4t kompensoida sit\u00e4 ep\u00e4j\u00e4rjest\u00e4ytyneill\u00e4 stressis\u00e4ikeill\u00e4. ZNF185:n on aikaisemmin liitetty eri sy\u00f6p\u00e4tyyppeihin, esim. keuhkosy\u00f6p\u00e4\u00e4n. Se voisikin mahdollisesti olla biomarkkeri, mink\u00e4 takia jatkossa onkin t\u00e4rke\u00e4\u00e4 tutkia tarkemmin ZNF185:n roolia stressis\u00e4iedynamiikoissa.", "language": "fi", "element": "description", "qualifier": "abstract", "schema": "dc"}, {"key": "dc.description.abstract", "value": "ZNF185 is one of the poorly characterized LIM-domain proteins with only 37 articles in Pubmed.org, just a few of them focusing on the cellular functions. LIM domain proteins typically facilitate protein-protein interactions that are involved in diverse cellular functions, which are often associated with the actin cytoskeleton. The role of ZNF185 was investigated further in this master\u2019s thesis by inactivating the ZNF185 gene in U-2 OS cells using CRISPR/Cas9-method. The successful knockout of ZNF185 was validated at the genomic as well as the protein level and all together five complete knockouts were identified. In addition to the complete knockout clones, some so-called partial knockout clones were identified that were expressing only smaller isoforms. Based on antiZNF185 stainings and exogenously expressed ZNF185 in U-2 OS wild-type cells, the subcellular localization of ZNF185 was determined mostly in the dorsal stress fibers and weakly to the ventral stress fibers. The immunofluorescence images of ZNF185 knockout clones revealed that the focal adhesions were elongated and larger compared to the wild-type cells. The dorsal stress fibers were faint or missing, which is in line with the subcellular localization of ZNF185. Moreover, the knockout cells formed tail structures which they could not retrack and they had irregular leading edge. This could indicate that there are cytoskeletal defects because of the loss of ZNF185 and the cell does not support the cell shape as it should. In the random cell migration assay, the speed of knockout cells was significantly slower, presumably due to the larger focal adhesions that take longer to disassemble. Nevertheless, the loss of ZNF185 had relatively small or non-significant effects on the directionality. The RhoA activity was measured in the knockout cells and wild-type cells to validate earlier studies, and it had no significant differences between the knockout cells and the wild-type. All in all, this data indicates that the ZNF185 deficient cells are under a lot of tension due to disordered cell cortex, and the knockout cells tried to compensate causing also the disheveled stress fibers. More studies must be done to research the specific role ZNF185 might have in the regulation of the actin cytoskeleton and focal adhesions because ZNF185 has a potential to be a biomarker for different types of cancer.", "language": "en", "element": "description", "qualifier": "abstract", "schema": "dc"}, {"key": "dc.description.provenance", "value": "Submitted by Miia Hakanen (mihakane@jyu.fi) on 2025-04-16T11:18:47Z\nNo. of bitstreams: 0", "language": "en", "element": "description", "qualifier": "provenance", "schema": "dc"}, {"key": "dc.description.provenance", "value": "Made available in DSpace on 2025-04-16T11:18:47Z (GMT). No. of bitstreams: 0\n Previous issue date: 2025", "language": "en", "element": "description", "qualifier": "provenance", "schema": "dc"}, {"key": "dc.format.extent", "value": "53", "language": null, "element": "format", "qualifier": "extent", "schema": "dc"}, {"key": "dc.language.iso", "value": "eng", "language": null, "element": "language", "qualifier": "iso", "schema": "dc"}, {"key": "dc.rights", "value": "In Copyright", "language": null, "element": "rights", "qualifier": null, "schema": "dc"}, {"key": "dc.subject.other", "value": "Actin filaments", "language": null, "element": "subject", "qualifier": "other", "schema": "dc"}, {"key": "dc.subject.other", "value": "CRISPR/Cas9", "language": null, "element": "subject", "qualifier": "other", "schema": "dc"}, {"key": "dc.subject.other", "value": "cytoskeleton", "language": null, "element": "subject", "qualifier": "other", "schema": "dc"}, {"key": "dc.subject.other", "value": "focal adhesion", "language": null, "element": "subject", "qualifier": "other", "schema": "dc"}, {"key": "dc.subject.other", "value": "LIM-domain", "language": null, "element": "subject", "qualifier": "other", "schema": "dc"}, {"key": "dc.subject.other", "value": "U-2 OS", "language": null, "element": "subject", "qualifier": "other", "schema": "dc"}, {"key": "dc.title", "value": "The role of ZNF185 in stress fibers", "language": null, "element": "title", "qualifier": null, "schema": "dc"}, {"key": "dc.type", "value": "master thesis", "language": null, "element": "type", "qualifier": null, "schema": "dc"}, {"key": "dc.identifier.urn", "value": "URN:NBN:fi:jyu-202504163305", "language": null, "element": "identifier", "qualifier": "urn", "schema": "dc"}, {"key": "dc.contributor.faculty", "value": "Matemaattis-luonnontieteellinen tiedekunta", "language": "fi", "element": "contributor", "qualifier": "faculty", "schema": "dc"}, {"key": "dc.contributor.faculty", "value": "Faculty of Sciences", "language": "en", "element": "contributor", "qualifier": "faculty", "schema": "dc"}, {"key": "dc.contributor.department", "value": "Bio- ja ymp\u00e4rist\u00f6tieteiden laitos", "language": "fi", "element": "contributor", "qualifier": "department", "schema": "dc"}, {"key": "dc.contributor.department", "value": "Department of Biological and Environmental Science", "language": "en", "element": "contributor", "qualifier": "department", "schema": "dc"}, {"key": "dc.contributor.organization", "value": "Jyv\u00e4skyl\u00e4n yliopisto", "language": null, "element": "contributor", "qualifier": "organization", "schema": "dc"}, {"key": "dc.contributor.organization", "value": "University of Jyv\u00e4skyl\u00e4", "language": null, "element": "contributor", "qualifier": "organization", "schema": "dc"}, {"key": "dc.subject.discipline", "value": "Solu- ja molekyylibiologia", "language": "fi", "element": "subject", "qualifier": "discipline", "schema": "dc"}, {"key": "dc.subject.discipline", "value": "Cell and molecular biology", "language": "en", "element": "subject", "qualifier": "discipline", "schema": "dc"}, {"key": "dc.type.coar", "value": "http://purl.org/coar/resource_type/c_bdcc", "language": null, "element": "type", "qualifier": "coar", "schema": "dc"}, {"key": "dc.rights.copyright", "value": "\u00a9 The Author(s)", "language": "fi", "element": "rights", "qualifier": "copyright", "schema": "dc"}, {"key": "dc.rights.accesslevel", "value": "restrictedAccess", "language": null, "element": "rights", "qualifier": "accesslevel", "schema": "dc"}, {"key": "dc.type.publication", "value": "masterThesis", "language": null, "element": "type", "qualifier": "publication", "schema": "dc"}, {"key": "dc.subject.yso", "value": "solut", "language": null, "element": "subject", "qualifier": "yso", "schema": "dc"}, {"key": "dc.rights.url", "value": "https://rightsstatements.org/page/InC/1.0/", "language": null, "element": "rights", "qualifier": "url", "schema": "dc"}, {"key": "dc.rights.accessrights", "value": "Tekij\u00e4 ei ole antanut lupaa avoimeen julkaisuun, joten aineisto on luettavissa vain Jyv\u00e4skyl\u00e4n yliopiston kirjaston arkistoty\u00f6semalta. Ks. https://kirjasto.jyu.fi/fi/tyoskentelytilat/laitteet-ja-tilat#autotoc-item-autotoc-2", "language": "fi", "element": "rights", "qualifier": "accessrights", "schema": "dc"}, {"key": "dc.rights.accessrights", "value": "The author has not given permission to make the work publicly available electronically. Therefore the material can be read only at the archival workstation at Jyv\u00e4skyl\u00e4 University Library (https://kirjasto.jyu.fi/fi/tyoskentelytilat/laitteet-ja-tilat#autotoc-item-autotoc-2.", "language": "en", "element": "rights", "qualifier": "accessrights", "schema": "dc"}, {"key": "dc.description.accessibilityfeature", "value": "unknown accessibility", "language": "en", "element": "description", "qualifier": "accessibilityfeature", "schema": "dc"}, {"key": "dc.description.accessibilityfeature", "value": "ei tietoa saavutettavuudesta", "language": "fi", "element": "description", "qualifier": "accessibilityfeature", "schema": "dc"}]
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