JNK, a versatile regulator of kinesin-1 transport and cytoskeleton dynamics

C-Jun N-terminal kinase (JNK) is traditionally recognized as a crucial factor in stress response and inducer of apoptosis upon various stimulations. Three isoforms build the JNK subfamily of MAPK; generally expressed JNK1 and JNK2 and brain specific JNK3. Degenerative potency placed JNK in the spotlight as potential pharmacological option for intervention. Unfortunately, adverse effects of potential drugs and observation that expression of only JNK2 and JNK3 are induced upon stress, restrained initial enthusiasm. Notably, JNK1 demonstrated atypical high constitutive activity in neurons that is not responsive to cellular stresses and indicated existence of physiological activity. This thesis aimed at revealing the physiological functions of JNK1 in actin homeostasis through novel effector MARCKS-Like 1 (MARCKSL1) protein, neuronal trafficking mediated by major kinesin-1 motor protein and microtubule (MT) dynamics via STMN2/SCG10. The screen for novel physiological JNK substrates revealed specific phosphorylation of C-terminal end of MARCKSL1 at S120, T148 and T183 both ex vivo and in vitro. By utilizing site-specific mutagenesis, various actin dynamics and migrations assays we were able to demonstrate that JNK1 phosphorylation specifically facilitates F-actin bundling and thus filament stabilisation. Consecutively, this molecular mechanism was proved to enhance formation of filopodia; cell surface projections that allow cell sensing surrounding environment and migrate efficiently. Our results visualize JNK dependent and MARCKSL1 executed induction of filopodia in neurons and fibroblast indicating general mechanism. Subsequently, inactivation of JNK action on MARCKSL1 shifts cellular actin machinery into lamellipodial dynamic arrangement. Tuning of actin cytoskeleton inevitably melds with cell migration. We observed that both active JNK and JNK pseudo-phosphorylated form of MARCKSL1 reduce actin turnover in intact cells leading to overall diminished cell motility. We demonstrate that tumour transformed cells from breast, prostate, lung and muscle-derived cancers upregulate MARCKSL1. We showed on the example of prostate cancer PC-3 cell line that JNK phosphorylation negatively controls MARCKSL1 ability to induce migration, which precedes cancer cell metastasis. The second round of identification of JNK physiological substrates resulted in detection of predominant motor protein kinesin-1 (Kif5). Mass spectrometry detailed analysis showed evident endogenous phosphorylation of kinesin-1 on S176 within motor domain that interacts with MT. In vitro phosphorylation of bacterially expressed kinesin heavy chain by JNK isoforms displayed higher specificity of JNK1 when compared to JNK3. Since, JNK1 is constitutively active in neurons it signified physiological aspect of kinesin-1 regulation. Subsequent biochemical examination revealed that kinesin-1, when not phosphorylated on JNK site, exhibits much higher affinity toward MTs. Expression of the JNK non-phosphorable kinesin-1 mutant in intact cells as well as in vitro single molecule imaging using total internal reflection fluorescence microscopy indicated that the mutant loses normal speed and is not able to move processively into proper cellular compartments. We identify novel kinesin-1 cargo protein STMN2/SCG10, which along with known kinesin-1 cargo BDNF is showing impaired trafficking when JNK activity is inhibited. Our data postulates that constitutive JNK activity in neurons is crucial for unperturbed physiologically relevant transport of kinesin-1 dependant cargo. Additionally, my work helps to validate another novel physiological JNK1 effector STMN2/SCG10 as determinant of axodendritic neurites dynamics in the developing brain through regulation of MT turnover. We show successively that this increased MT dynamics is crucial during developmental radial migration when brain layering occurs. Successively, we are able to show that introduction of JNK phosphorylation mimicking STMN2/SCG10 S62/73D mutant rescues completely JNK1 genetic deletion migration phenotype. We prove that STMN2/SCG10 is predominant JNK effector responsible for MT depolymerising activity and neurite length during brain development. Summarizing, this work describes identification of three novel JNK substrates MARCKSL1, kinesin-1 and STMN2/SCG10 and investigation of their roles in cytoskeleton dynamics and cargo transport. This data is of high importance to understand physiological meaning of JNK activity, which might have an adverse effect during pharmaceutical intervention aiming at blocking pathological JNK action.

Stretching a service concept brand

Brands have become one of the most valuable assets for organizations, which is why organizations want to benefit from them in whole scale. However, service concept brands are still little researched. The purpose of this research is to study how brand stretching and co-branding strategies can help a service concept brand to attain wider target market. More specifically, this research aims to contribute the methods and their risks and benefits to different customer levels of a group fitness brand. This study is a qualitative single case analysis embedded with multiple units of analysis. The data used in this study was gathered by nine theme interviews. The interviewees are from one of the customer levels of the service concept provided by the case organization. The interviews are made in different geographical areas in Finland. The results of the study will clarify and illustrate the differences and similarities between the theoretical framework and practise. Several differences between traditional brand stretching and co-branding strategies and those that are possible to employ by a service concept brand were found. The answers of the interviewees were slightly different depending on their role in the organization and their experience from the branch. However, they proved that not all brand stretching or co-branding strategies are applicable in the group fitness brand. Nevertheless, also several similarities that benefit the group fitness brand were found.

Exploring the value creation in service ecosystem context: the case of the information technology sector

This thesis examines customer value creation in a service ecosystem context. The objective of this thesis is to develop a comprehensive view of value creation processes in a service ecosystem context and an understanding on the roles of the stakeholders involved in these processes, focusing on the information technology industry. The novelty of the two central concepts of this thesis, systemic customer value and service ecosystem, as well as the gap in the literature of empirical research on value creation in an ecosystem-level, opened an interesting research topic. The empirical study is conducted as a single case analysis, utilizing Group Decision Support System (GDSS) and also Analytic Hierarchy Process (AHP). The findings suggest that customer value is created by a complex combination of interactions among different actors of the ecosystem. Thus, value is not created by a single offering directed to the customer, but by an integration of services from different parts of the ecosystem as well as the active participation of customer in this process.

Tuotteiston nykytila-analyysi ja kehitystoimenpiteiden kartoitus ohutlevytuotteita valmistavassa yrityksessä

Työn tavoitteena oli tutkia kohdeyrityksen tuotteiston nykytilaa ja antaa kehitysehdotus tuotteiston uudesta rakenteesta, jolla voidaan toteuttaa tehokasta sarjatuotantoa. Tutkimuksessa keskityttiin vakiotuotteista koostuvan tuotteiston analysointiin. Tarkastelun kohteeksi valittiin tuotteiston yksittäiset tuotenimikkeet, joita analysoitiin toteutuneiden myyntilukujen ja asiakashaastatteluista saatujen tietojen perusteella Työssä käytettiin tutkimusmenetelminä kirjallisuustutkimusta, teemahaastattelua ja tuotteiden myynti- ja käyttötilastojen tarkastelua. Tutkimuksen luotettavuutta lisäsi, kun käytettiin kolmen riippumattoman tutkimusta vahvistavan menetelmän yhdistämistä. Tutkimuksessa saavutettujen tulosten perusteella tuotteille tulisi kehittää uudet ohjausmenetelmät, jotka helpottavat tuotannon ja varaston hallintaa. Tuotteet, joilla ei ole ollut myyntiä, tulisi käydä yksityiskohtaisesti läpi ja tehdä arviointi niiden tarpeellisuudesta. Tuoterakenteita tulisi kehittää modulaarisemmaksi tehokkaampaa sarjatuotantoa ajatellen. Tuoteryhmissä olevat samankaltaiset tuotteet tulisi yhdistää, jolloin valikoimasta saataisiin yksinkertaisempi ja helpommin hallittava.

Customer References as Marketing Practice in Company Web sites - Content and Discourse Analysis

Työn tarkoituksena oli tutkia sisältö- ja diskurssianalyysin avulla kuinka yritykset viestivät asiakasreferenssejä verkkosivuillaan. Työssä keskityttiin tutkimaan yritysten referenssikuvausten teemoja ja diskursseja, sekä sitä kuinka referenssisuhde rakentuu diskursiivisesti referenssikuvauksissa. Tutkimukseen valittiin kolme suomalaista ICT-alan yritystä: Nokia, TietoEnator ja F-Secure. Aineisto koostuu 140:stä yritysten WWW-sivuilta kerätystä referenssikuvauksesta. Sisältöanalyysin tuloksena havaittiin, että referenssikuvaukset keskittyvät kuvaamaan yksittäisiä tuote- tai projektitoimituksia referenssiasiakkaille kyseisten asiakassuhteiden valossa. Analyysin tuloksena tunnistettiin kolme diskurssia: hyötydiskurssi, sitoutumisen diskurssi sekä teknologisen eksperttiyden diskurssi. Diskurssit paljastavat referenssikuvausten retoriset keinot ja konstruoivat referenssisuhteen ja toimittajan subjektiposition eri näkökulmista. Pääpaino referenssikuvauksissa on toimittajan ratkaisun tuomissa hyödyissä. Diskurssit tuottavat referenssisuhteesta kuvan hyötyjä tuovana ja läheisenä asiakassuhteena, joka tarjoaa väylän ulkopuolisiin kyvykkyyksiin ja teknologioihin. Toimittaja esitetään referenssikuvauksissa diskurssista riippuen hyötyjen tuojana, luotettavana partnerina sekä kokeneena eksperttinä. Referenssiasiakas sen sijaan esitetään vain yhdestä näkökulmasta stereotyyppisesti tärkeänä ja tyytyväisenä asiakkaana.