13 research outputs found
Prof. N. NAGARAJ
No full textUniversity of Agricultural Sciences, Department of Agricultural Economics, Agriculture College, Bangalore, Karnataka Thème de recherche : Connecting, Enhancing and Sustaining Environmental Services and Market Values of Coffee, Agro Foresty in Indi
Prof. N. NAGARAJ
No full textUniversity of Agricultural Sciences, Department of Agricultural Economics, Agriculture College, Bangalore, Karnataka Thème de recherche : Connecting, Enhancing and Sustaining Environmental Services and Market Values of Coffee, Agro Foresty in Indi
Do Labor Intensive Industries Generate Employment? Evidence from firm level survey in India
Get PDFThis study attempts to address the issue of declining labour intensity in Indias organized manufacturing in order to understand the constraints on employment generation in the labour intensive sectors. Using primary survey data covering 252 labour intensive manufacturing-exporting firms across five sectorsapparel, leather, gems and jewellery, sports goods, and bicycles for 2005-06 an attempt is made to find out the factors which constrain employment generation in labour intensive firms. The study shows several constraints in the path of employment generation in labour intensive sectorsnon-availability of trained skilled workers, infrastructure bottlenecks, low levels of investment, labour rules and regulations, and a noncompetitive export orientation. The study suggests a set of policy initiatives to improve the employment potential of these sectors.Indian Organized Manufacturing, Labor Intensity, Employment Growth, Skilled workforce, Wage Structure, Export status, Machinery Usage, Labor laws, South Asia
Phospho-regulation of the Shugoshin-Condensin interaction at the centromere in budding yeast
Get PDFAuthor summary Proper chromosome segregation in eukaryotes is ensured through correct attachment of the spindle microtubules to the centromeric chromosomal regions. The attachment is mediated via the multimolecular proteinaceous complex called the kinetochore. This enables the establishment of bioirentation, when each sister chromatid is attached to microtubules emanating from opposite spindle poles. Shugoshin (Sgo1) is a conserved centromeric protein that facilitates biorientation through its interactions with the protein phosphatase PP2A/Rts1, chromosome passenger complex and centromeric condensin. Here, we identified a serine-rich motif that is required for the interaction of shugoshin with the condensin complex. We show that loss of this region impairs condensin enrichment at the centromere, chromosome biorientation, segregation as well as the function of the chromosome passenger complex in the error correction. Moreover, the interaction is phosphoregulated, as phosphorylation of the serine-rich motif on Sgo1 disrupts its interaction with condensin. Finally, we show that the conserved spindle assembly checkpoint kinase Mps1 is responsible for this phosphorylation. Our findings uncover novel regulatory mechanisms that facilitate proper chromosome segregation. Correct bioriented attachment of sister chromatids to the mitotic spindle is essential for chromosome segregation. In budding yeast, the conserved protein shugoshin (Sgo1) contributes to biorientation by recruiting the protein phosphatase PP2A-Rts1 and the condensin complex to centromeres. Using peptide prints, we identified a Serine-Rich Motif (SRM) of Sgo1 that mediates the interaction with condensin and is essential for centromeric condensin recruitment and the establishment of biorientation. We show that the interaction is regulated via phosphorylation within the SRM and we determined the phospho-sites using mass spectrometry. Analysis of the phosphomimic and phosphoresistant mutants revealed that SRM phosphorylation disrupts the shugoshin-condensin interaction. We present evidence that Mps1, a central kinase in the spindle assembly checkpoint, directly phosphorylates Sgo1 within the SRM to regulate the interaction with condensin and thereby condensin localization to centromeres. Our findings identify novel mechanisms that control shugoshin activity at the centromere in budding yeast.We thank Nagarunja Nagaraj for his help with the identification of the phosphosites on Sgo1 that was performed by the Mass Spectrometry Core Facility at the MPI Biochemistry, Martinsried, Germany
Polarization, inequality and growth: The Indian experience
Get PDFWe analyze polarization in India roughly in the past two and half decades using consumption expenditure data. We show that both bipolarization and multidimensional polarization (on several dimensions: caste, rural-urban, state, region) have increased since the 1990s. In the case of bipolarization, this is a reversal from the earlier trend (in the 1980s). Overall, our results suggest that the high growth that India has been witnessing since the 1990s has been associated with widening disparities. Comparing polarization and inequality, we find similarities, but also some differences. Our results therefore underscore the importance of studying polarization as distinct from traditional inequality.Polarization; Inequality; Growth in India.
Proteomic investigation of the class IA phosphoinositide 3-kinase signalling pathway
No full textPhDClass IA phosphoinositide 3-kinases (PI3Ks) are a family of enzymes with key roles in the regulation of signalling pathways, many of which are mediated through Akt. The PI3K/Akt pathway has a critical role in the regulation of key cellular functions such as cell survival, growth, proliferation, migration and metabolism and is involved in several diseases.
Advances in mass spectrometry and separation sciences have revolutionized the field of proteomics. Techniques based on mass spectrometry are now the first choice for protein identification and are increasingly important for their quantitative analysis. The development of phosphoproteomics has also permitted the global quantitative analysis of signalling activity.
The aim of this project was to contribute to our understanding of the activity and regulation of the PI3K/Akt signalling pathway from a proteomics perspective. Novel approaches based on mass spectrometry were designed, developed and applied to the investigation of this signalling pathway.
Firstly, we investigated the hypothesis that PI3K activity may be regulated by dynamic protein interactions. We designed an affinity purification mass spectrometry strategy to identify proteins interacting dynamically with PI3K. Our study revealed that calpain small subunit 1 interacts dynamically with PI3K. Further investigation demonstrated that active calpain heterodimers associate dynamically with PI3K, thereby regulating PI3K stability and activity.
Secondly, we characterized phosphorylation events downstream of Akt, a major downstream effector of PI3K. We used global shotgun phosphoproteomics of a cell line expressing an inducible construct encoding constitutively active Akt to identify phosphorylation events downstream of Akt in vivo. In addition, we also developed an in vitro kinase assay which, when coupled to global shotgun phosphoproteomics, enables the quantification of Akt1 activity in addition to the identification of downstream phosphorylation sites. Furthermore, we found that this in vitro approach may be used as a method suitable for the global profiling of endogenous kinase activities.Medical Research Counci
Employee Engagement and Relationship Practices in Start-up organizations
Get PDFIn order to stay competitive in today’s fast changing world, presence of the best talent depicts the success or failure of the organisation. Who you hire is not just important, but how the employee is managed and retained is considered important by Kennedy and Diam, 2010. Initially considered as a cheap labour force provider, India witnessed reduction in product development and innovation. By the introduction of initiatives by the new Government, raise in funding of about 5 billion in 2015 marked the beginning of job providing market in India. Hiring and retaining skilled manpower is considered as a huge challenge for the start-up to be successful and uncertain organisation climate change is the reason. (Banerjee 2017, Lee Hom and Liz 2017). The inherent uncertainty of organizational growth has resulted in a level of systematic flexibility regarding internal operations requiring high endand allowing for contingent leadership to take place. While flexibility allows optimal resource utilization, on the other hand it can also result in employees feeling aloof, stressed and de-motivated (Banerjee 2017).If these issues remain unanswered, the employee becomes disengaged and leaves for better opportunity. This study focuses on HRM practices (i.e. employee engagement and retention) in start-ups, effectiveness of practices (employee satisfaction) and challenges the start-ups will face in competitive setup. The study makes use of qualitative data and is collected through narrative enquiry. The data isanalysed using ISM technique; excel with focus on HRM practices start-ups and its impact on retentionship and engagement outcomes
The TRF1 telomere protein is essential for the generation and maintenance of iPS cells and marks both pluripotent and adult stem cells
Get PDFAbstract Englisch: Telomeres are nucleoprotein structures that protect the chromosomal ends from being recognized by DNA repair mechanisms as DNA double strand breaks. The Telomeric DNA is bound by various proteins that force the whole structure to fold in the so-called telomeric loop, hiding the DNA ends in the double stranded DNA. One of those sheltering proteins is the telomere repeat binding factor 1 (TRF1) which binds to the double-stranded telomeric DNA and is implicated in telomere length regulation. TRF1-deficient mice are embryonic lethal at the blastocyst stage and the conditional deletion of TRF1 in stratified epithelia leads to hair follicle stem cell defects, suggesting thus a role for TRF1 in stemness.
To address this and also the possibility of using TRF1 as an in vivo telomere length marker, here we generated a reporter mouse carrying a knock-in (KI) allele in which TRF1 is fused to the reporter protein eGFP. We find that eGFP-TRF1 expression is maximal at the adult stem cell compartments in the mouse, including the hair follicle stem cell niche and Lgr5-positive (Lgr5+) and Lgr5-low/negative stem cells at the intestinal crypts, and that eGFP-TRF1 expression is uncoupled from telomere length. Conditional deletion of TRF1 in the small intestine leads to a rapid collapse of the villi/crypt structures coincidental with increased DNA damage and apoptosis, indicating that TRF1 is essential to maintain small intestine homeostasis. Thus, TRF1 both marks adult stem cell compartments and is essential for their functionality. In line with this, we found very high levels of eGFP-TRF1 in induced pluripotent stem (iPS) cells, levels that are uncoupled from telomere elongation associated to reprogramming. TRF1 in iPS cells is expressed heterogeneous and coincident with the in-built heterogeneity of Nanog expression in iPS cell colonies. Selection of high eGFP-TRF1 iPS cells correlated with a higher pluripotency as indicated by their ability to form teratomas and chimeras. By using various loss-of-function approaches, we show that TRF1 is necessary for both the induction and the maintenance of pluripotency, by preventing the induction of DNA damage response and apoptosis. Finally, supporting the notion that TRF1 is a key factor for pluripotency we make the unprecedented finding that TRF1 is a direct target of the transcription factor Oct3/4, which binds to the TRF1 promoter and increases the transcription of TRF1, thus providing a mechanistic link between TRF1 and pluripotency.
These findings render TRF1 a novel marker for stem cells, those cells having the highest abundance of TRF1 in a given tissue. Also they deploy the eGFP-TRF1 mouse model as a useful tool for following up such stem cells in vivo and in vitro. ---------- Abstract Deutsch: Werden lose Chromosomenenden von DNS-Reparaturmechanismen erkannt führt dies zu Chromosomenfusionen, Zellseneszenz und Apoptose. Telomere, die Endstücke chromosomaler DNS sind Nukleoprotein-Strukturen mit der Aufgabe die Chromosomenenden davor zu schützen. Die telomerische DNS wird dabei von verschiedenen Proteinen erkannt, gebunden und zu einer Schleife geformt, deren Ende in der telomer-duplex DNA versteckt wird. Eines dieser Schutzproteine ist der „Telomere repeat binding factor 1“ (TRF1), welches die doppelsträngige Telomer-DNS bindet und bei der Regulation der Telomerlänge beteiligt ist. Wird TRF1 depletiert, sterben Embryonen bereits als Blastozysten, zeigen aber keine Telomeranomalitäten. Das konditionelle Ausschalten von TRF1 im mehrschichtigen Epithel (Keratin 5 exprimierendes Epithel) führt jedoch zu hoher Telomerfragilität und zum Verlust von Haarfollikeln was auf einen Stammzelldefekt hindeutet.
Um mögliche Stammzelleigenschaften sowie die Möglichkeit von TRF1 als in vivo Telomerlängenindikator zu eruieren, wurde hier eine knock-in Maus generiert, bei der eine eGFP-Kassette in das Aminoende des TRF1-Lokus fusioniert wurde. Mit diesem Model konnte ich zeigen, dass die eGFP-TRF1-Expression in den adulten Stammzellnischen der Haarfollikel und der Dünndarm-Krypten erhöht ist. Konditionelles ausknocken von TRF1 im Dünndarm führte zu einem schnellen Kollaps der Villi/Krypt-Struktur sobald TRF1 in allen Stammzellen eliminiert war. Übereinstimmend mit den erhöhten TRF1-Levels in adulten Stammzellen, zeigten auch induzierte, pluripotente Stammzellen (iPSz) eine deutlich erhöhte Expression von eGFP-TRF1, was aufgrund aktiver Telomerase und sich verlängernder Telomere erwartet werden konnte. Überraschenderweise ist die Menge an TRF1 während des Reprogramierungsprozesses überproportional zur Telomerlänge angestiegen; auch in Zellen die keine Fähigkeit hatten die Telomere zu verlängern. Die Expression von eGFP-TRF1 in iPS-Zellkolonien ist heterogen und korreliert mit Nanog, einem Stammzellmarker der in iPS-Zellen ebenfalls variiert und die Pluripotenz der Zellen positiv beeinflusst. IPS-Zellen mit hoher eGFP-TRF1 Expression waren potenter, subkutane Teratome zu bilden und beteiligten sich an der Formation von Chimären. Mit verschiedenen TRF1 knock-out und knock-down Studien konnte ich belegen, dass TRF1 für die Induktion, wie auch für die Aufrechterhaltung der Pluripotenz essentiell ist. Ebenfalls bewiesen wir, dass OCT3/4, ein Transkriptionsfaktor für Pluripotenzgene, an die TRF1 Promoter-Region bindet und die TRF1 Transkription positiv beeinflusst.
Diese Resultate zeigen TRF1 als einen wesentlichen Faktor der für Pluripotenz und einem neuen Marker für Stammzellen in vitro und in vivo. EGFP-TRF1 konnte aber nicht als in vivo Telomerlängenindikator etabliert werden. ---------- Abstract Espanol: Los telómeros son estructuras nucleoproteicas que evitan que los extremos de los cromosomas sean reconocidos por los mecanismos de reparación del ADN como roturas en la doble hebra de ADN. El ADN telomérico está unido a varias proteínas que facilitan la formación del denominado bucle telomérico, y que protege los extremos del ADN bicatenario. La proteína TRF1 (telomere repeat binding factor 1) es un miembro del complejo proteíco especializado de protección telomérica que se une al ADN cadena doble y está implicada en la regulación de la longitud telomérica. Los ratones deficientes en TRF1 adolecen de letalidad embrionaria en la etapa de blastocito y la deleción condicional de esta proteína en el epitelio estratificado conduce a defectos en las células madre del folículo piloso, lo que sugiere por tanto una implicación funcional de TRF1 en pluripotencia.
Con el fin de elucidar esta conexión entre TRF1 y pluripotencia y también el posible empleo de TRF1 como un marcador de longitud telomérica in vivo, generamos un ratón “reporter” que contiene un alelo knock-in (KI) en el que TRF1 está fusionada a la proteína eGFP. La expresión de eGFP-TRF1 alcanza, de manera independiente de la longitud telomérica, valores máximos en los compartimentos de células madre adultas, entre los que se incluyen el nicho de células madre del folículo piloso y las células madre Lgr5-positivas (Lgr5+) y Lgr5-negativas (Lgr5-) de las criptas intestinales. La deleción condicional de TRF1 en el intestino delgado desencadena un rápido colapso de las vellosidades y criptas intestinales, coincidente con un aumento en el daño en el ADN y en la apoptosis, lo cual indica que TRF1 es esencial en el mantenimiento de la homeostasis del intestino delgado. Por consiguiente, TRF1 demarca los compartimentos de células madre adultas y es indispensable para su funcionalidad. En consonancia con lo anterior, describimos la presencia de niveles muy elevados de eGFP-TRF1 en las células madre pluripotentes inducidas (células iPS), valores que son independientes del alargamiento telomérico asociado a la reprogramación nuclear. Además, los niveles de TRF1 entre las diferentes células de las colonias de Ips son heterogéneos y coincidentes con la también intrínsecamente heterogénea expresión de Nanog. En las células iPS los altos niveles de eGFP-TRF1 correlacionan con una mayor pluripotencia, asociada a una mayor capacidad para formar teratomas y quimeras. Por medio de diferentes ensayos de pérdida de función, demostramos que TRF1 es necesaria para la inducción y mantenimiento de la pluripotencia, evitando que se desencadene la respuesta de daño en el ADN y la apoptosis. Por último, la idea de que TRF1 sea un factor clave se ve respaldada por el hallazgo sin precedentes de que TRF1 es una diana directa del factor de transcripción Oct3/4, cuya unión al promotor de TRF1 activa la transcripción de TRF1, lo que proporciona un mecanismo que conecta la proteína telomérica TRF1 con la pluripotencia.
Estos resultados convierten a TRF1 en un nuevo marcador de células madre, debido a que su expresión es máxima en las células madre de un tejido dado y al modelo de ratón eGFP-TRF1 en una útil herramienta para el seguimiento de estas células madre in vivo e in vitro
The molecular dysregulation of excitation contraction coupling in patients with congenital muscle disorders
Get PDFExcitation contraction coupling (ECC) is the process whereby an action potential spreading throughout the muscle membrane activates muscle contraction, by releasing Ca2+ from the Sarcoplasmic Reticulum (SR). Ca2+ release from the SR is mediated by the Ryanodine Receptor located on the SR membrane. Any alterations in the architecture of the intercellular muscle membrane compartments or mutations in the RYR1 gene are associated with neuromuscular disorders such as Central core disease, Multi minicore disease, Central nuclear myopathy or congenital fiber type disproportion.
In the last few decades, ECC characteristics were extensively investigated in our lab, on myotubes originating from patient’s muscle satellite cells. In the 1st paper entitled “Establishment of human skeletal muscle- derived cell line: biochemical, cellular and electrophysiological characterization”, we studied the ECC in an immortalized human muscle cell line (HMCL-7304), which helps to overcome many of the technical limitations of working with primary muscle cells from human patients. ECC in HMCL-7304 was characterized with qPCR and western blotting as well as super resolution microscopy (SIM), Ca2+ imaging and electrophysiological measurements. We discovered that HMCL-7304 have a phenotype closer to slow twitch muscles than fast twitch muscles. HMCL-7304 can be used as a platform to investigate genetic mechanisms of muscle disorders, as shown in our 2nd publication; “RyR1 deficiency in congenital myopathies disrupts excitation contraction coupling”, where we simulated the downregulation of RyR1 expression as seen in patients with recessive RYR1 mutations, by silencing RyR1 expression in the HMCL-7304.
Patients with recessive RYR1 mutations have been shown to downregulate RyR1 expression in skeletal muscles. This is in contrast to what is observed in patients with dominant RYR1 mutations, in whom we could not find reduction in the RyR1 expression. In patient’s muscle biopsies where RyR1 expression is reduced, all isoforms of InsP3R Receptors (ITPR1-ITPR3) were found to be up-regulated. Ca2+ release was not altered by the reduction of RyR1 expression using siRNA in HMCL or by blocking of IP3Rs using Xestospongin, rejecting the possibility for InsP3R functional compensation for the downregulation of RyR1.
The potential mechanisms causing downregulation of RyR1 in patients with recessive RYR1 mutations is addressed in our 3rd publication; “Epigenetic changes as a common trigger of muscle weakness in congenital myopathies”. Patients with downregulation of RyR1, exhibit decreased expression of muscle specific microRNAs and increased expression of HDAC4 and HDAC5. Additionally hyper-methylation of CpG Island in the RYR1 gene was observed. Down regulation of RyR1, downregulation microRNAs and upregulation of HDAC4 and HDAC5 was also observed in patients with Nemaline myopathy, reflecting common epigenetic changes activated in congenital myopathies. Using HDAC or DNMT inhibitors can target common downstream pathways activated in muscles of patients with congenital myopathies offers an interesting new approach for the amelioration of muscle functio
