Transcriptional and translational regulators of gene expression in germ cell development

Germ cell development is a highly coordinated process driven, in part, by regulatory mechanisms that control gene expression. Not only transcription, but also translation, is under regulatory control to direct proper germ cell development. In this dissertation, I have focused on two regulators of germ cell development. One is the homeobox protein RHOX10, which has the potential to be both a transcriptional and translational regulator in mouse male germ cell development. The other is the RNA-binding protein, Hermes, which functions as a translational regulator in Xenopus laevis female germ cell development. ^ Rhox10 is a member of reproductive homeobox gene X-(linked (Rhox) gene cluster, of which expression is developmentally regulated in developing mouse testes. To identify the cell types and developmental stages in which Rhox10 might function, I characterized its temporal and spatial expression pattern in mouse embryonic, neonatal, and adult tissues. Among other things, this analysis revealed that both the level and the subcellular localization of RHOX10 are regulated during germ cell development. To understand the role of Rhox10 in germ cell development, I generated transgenic mice expressing an artificial microRNA (miRNA) targeting Rhox10. While this artificial miRNA robustly downregulated RHOX10 protein expression in vitro, it did not significantly reduce RHOX10 expression in vivo. So I next elected to knockdown RHOX10 levels in spermatogonial stem cells (SSCs), which I found highly express both Rhox10 mRNA and RHOX10 protein. Using a recently developed in vitro culture system for SSCs combined with a short-hairpin RNA (shRNA) approach, I strongly depleted RHOX10 expression in SSCs. These RHOX10-depleted cells exhibited a defect in the ability to form stem cell clusters in vitro. Expression profiling analysis revealed many genes regulated by Rhox10, including many meiotic genes, which could be downstream of Rhox10 in a molecular pathway that controls SSC differentiation. ^ RNA recognition motif (RRM) containing protein, Hermes is localized in germ plasm, where dormant mRNAs are also located, of Xenopus oocytes, which implicates its role in translational regulator. To understand the function of Hermes in oocyte meiosis, I used a morpholino oligonucleotide (MO) based knockdown approach. Microinjection of Hermes MO into fully grown oocytes, which are arrested in meiotic prophase, caused acceleration of oocytes reentry into meiosis (i.e., maturation) upon progesterone induction. Using a candidate approach, I identified at least three targets of Hermes: Ringo/Spy, Xcat2, and Mos. Ringo/Spy and Mos are known to have functions in oocyte maturation, while Ringo/Spy, Xcat2 mRNA are localized in the germ plasm of oocytes, which drives germ cell specification after fertilization. This led me to propose that Hermes functions in both oocyte maturation and germ cell development through its ability to regulate 3 crucial target mRNAs. ^

Palynology on sediment profile Tso Kar in Ladakh

Palynological investigation of a 410 cm long core section from Tso Kar (33°10'N, 78°E, 4527 m a.s.l.), an alpine lake situated in the arid Ladakh area of NW India at the limit of the present-day Indian summer monsoon, was performed in order to reconstruct post-glacial regional vegetation and climate dynamics. The area was covered with alpine desert vegetation from ca. 15.2 to 14 kyr BP (1 kyr=1000 cal. years), reflecting dry and cold conditions. High influx values of long-distance transported Pinus sylvestris type pollen suggest prevailing air flow from the west and northwest. The spread of alpine meadow communities and local aquatic vegetation is a weak sign of climate amelioration after ca. 14 kyr BP. Pollen data (e.g. influx values of Pinus roxburghii type and Quercus) suggest that this was due to a strengthening of the summer monsoon and the reduced activity of westerly winds. The further spread of Artemisia and species-rich meadows occurred in response to improved moisture conditions between ca. 12.9 and 12.5 kyr BP. The subsequent change towards drier desert-steppe vegetation likely indicates more frequent westerly disturbances and associated snowfalls, which favoured the persistence of alpine meadows on edaphically moist sites. The spread of Chenopodiaceae-dominated vegetation associated with an extremely weak monsoon occurred at ca. 12.2-11.8 kyr BP during the Younger Dryas interstadial. A major increase in humidity is inferred from the development of Artemisia-dominated steppe and wet alpine meadows with Gentianaceae after the late glacial/early Holocene transition in response to the strengthening of the summer monsoon. Monsoonal influence reached maximum activity in the Tso Kar region between ca. 10.9 and 9.2 kyr BP. The subsequent development of the alpine meadow, steppe and desert-steppe vegetation points to a moderate reduction in the moisture supply, which can be linked to the weaker summer monsoon and the accompanying enhancement of the winter westerly flow from ca. 9.2 to 4.8 kyr BP. The highest water levels of Tso Kar around 8 kyr BP probably reflect combined effect of both monsoonal and westerly influence in the region. An abrupt shift towards aridity in the Tso Kar region occurred after ca. 4.8 kyr BP, as evidenced by an expansion of Chenopodiaceae-dominated desert-steppe. Low pollen influx values registered ca. 2.8-1.3 kyr BP suggest scarce vegetation cover and unfavourable growing conditions likely associated with a further weakening of the Indian Monsoon.

Pollen and non-pollen palynomorph records of the Holocene part of the composite sediment core TMD from lake Tso Moriri (NW India) and of modern surface samples from the Tso Moriri region

The high-altitude lake Tso Moriri (32°55'46'' N, 78°19'24'' E; 4522 m a.s.l.) is situated at the margin of the ISM and westerly influences in the Trans-Himalayan region of Ladakh. Human settlements are rare and domestic and wild animals are concentrating at the alpine meadows. A set of modern surface samples and fossil pollen from deep-water TMD core was evaluated with a focus on indicator types revealing human impact, grazing activities and lake system development during the last ca. 12 cal ka BP. Furthermore, the non-pollen palynomorph (NPP) record, comprising remains of limnic algae and invertebrates as well as fungal spores and charred plant tissue fragments, were examined in order to attest palaeolimnic phases and human impact, respectively. Changes in the early and middle Holocene limnic environment are mainly influenced by regional climatic conditions and glacier-fed meltwater flow in the catchment area. The NPP record indicates low lake productivity with high influx of freshwater between ca. 11.5 and 4.5 cal ka BP which is in agreement with the regional monsoon dynamics and published climate reconstructions. Geomorphologic observations suggest that during this period of enhanced precipitation the lake had a regular outflow and contributed large amounts of water to the Sutlej River, the lower reaches of which were integral part of the Indus Civilization area. The inferred minimum fresh water input and maximum lake productivity between ca. 4.5-1.8 cal ka BP coincides with the reconstruction of greatest aridity and glaciation in the Korzong valley resulting in significantly reduced or even ceased outflow. We suggest that lowered lake levels and river discharge on a larger regional scale may have caused irrigation problems and harvest losses in the Indus valley and lowlands occupied by sedentary agricultural communities. This scenario, in turn, supports the theory that, Mature Harappan urbanism (ca. 4.5-3.9 cal ka BP) emerged in order to facilitate storage, protection, administration, and redistribution of crop yields and secondly, the eventual collapse of the Harappan Culture (ca. 3.5-3 cal ka BP) was promoted by prolonged aridity. There is no clear evidence for human impact around Tso Moriri prior to ca. 3.7 cal ka BP, with a more distinct record since ca. 2.7 cal ka BP. This suggests that the sedimentary record from Tso Moriri primarily archives the regional climate history.

Extragenic Suppressors of the Arabidopsis gai Mutation Alter the Dose-Response Relationship of Diverse Gibberellin Responses1

Active gibberellins (GAs) are endogenous factors that regulate plant growth and development in a dose-dependent fashion. Mutant plants that are GA deficient, or exhibit reduced GA responses, display a characteristic dwarf phenotype. Extragenic suppressor analysis has resulted in the isolation of Arabidopsis mutations, which partially suppress the dwarf phenotype conferred by GA deficiency and reduced GA-response mutations. Here we describe detailed studies of the effects of two of these suppressors, spy-7 and gar2–1, on several different GA-responsive growth processes (seed germination, vegetative growth, stem elongation, chlorophyll accumulation, and flowering) and on the in planta amounts of active and inactive GA species. The results of these experiments show that spy-7 and gar2–1 affect the GA dose-response relationship for a wide range of GA responses and suggest that all GA-regulated processes are controlled through a negatively acting GA-signaling pathway.

Gibberellin Dose-Response Regulation of GA4 Gene Transcript Levels in Arabidopsis1

The gibberellins (GAs) are a complex family of diterpenoid compounds, some of which are potent endogenous regulators of plant growth. As part of a feedback control of endogenous GA levels, active GAs negatively regulate the abundance of mRNA transcripts encoding GA biosynthesis enzymes. For example, Arabidopsis GA4 gene transcripts encode GA 3β-hydroxylase, an enzyme that catalyzes the conversion of inactive to active GAs. Here we show that active GAs regulate GA4 transcript abundance in a dose-dependent manner, and that down-regulation of GA4 transcript abundance is effected by GA4 (the product of 3β-hydroxylation) but not by its immediate precursor GA9 (the substrate). Comparison of several different GA structures showed that GAs active in promoting hypocotyl elongation were also active in regulating GA4 transcript abundance, suggesting that similar GA:receptor and subsequent signal transduction processes control these two responses. It is interesting that these activities were not restricted to 3β-hydroxylated GAs, being also exhibited by structures that were not 3β-hydroxylated but that had another electronegative group at C-3. We also show that GA-mediated control of GA4 transcript abundance is disrupted in the GA-response mutants gai and spy-5. These observations define a sensitive homeostatic mechanism whereby plants may regulate their endogenous GA levels.

Gibberellins Promote Trichome Formation by Up-Regulating GLABROUS1 in Arabidopsis1

Trichome development is dependent on gibberellin (GA) signaling in Arabidopsis thaliana. Using the GA-deficient mutant ga1–3, the GA-response mutant spy-5, and uniconazol (a GA-biosynthesis inhibitor), we show that the GA level response correlates positively with both trichome number and trichome branch number. Two genes, GL1 and TTG, are required for trichome initiation. In ga1–3, coexpression of GL1 and R, the maize TTG functional homolog, under control of the constitutive 35S promoter, restored trichome development, whereas overexpression of neither GL1 nor R alone was sufficient to significantly suppress the glabrous phenotype. We next focused on GL1 regulation by GAs. In the double mutant the gl1–1 glabrous phenotype is epistatic to the spy-5 phenotype, suggesting that GL1 acts downstream of the GA signal transduction pathway. The activity of a β-glucuronidase reporter gene driven by the GL1 promoter was decreased in the wild type grown on uniconazol and showed a clear GA-dependent activation in ga1–3. Finally, quantification of GL1 transcript levels by reverse transcriptase-polymerase chain reaction demonstrated that relative to wild type, ga1–3 plants contained less transcript. These data support the hypothesis that GAs induce trichome development through up-regulation of GL1 and possibly TTG genes.

SPINDLY, a tetratricopeptide repeat protein involved in gibberellin signal transduction in Arabidopsis.

Gibberellins (GAs) are a major class of plant hormones that control many developmental processes, including seed development and germination, flower and fruit development, and flowering time. Genetic studies with Arabidopsis thaliana have identified two genes involved in GA perception or signal transduction. A semidominant mutation at the GIBBERELLIN INSENSITIVE (GAI) locus results in plants resembling GA-deficient mutants but exhibiting reduced sensitivity to GA. Recessive mutations at the SPINDLY (SPY) locus cause a phenotype that is consistent with constitutive activation of GA signal transduction. Here we show that a strong allele of spy is completely epistatic to gai, indicating that SPY acts downstream of GAI. We have cloned the SPY gene and shown that it encodes a new type of signal transduction protein, which contains a tetratricopeptide repeat region, likely serving as a protein interaction domain, and a novel C-terminal region. Mutations in both domains increase GA signal transduction. The presence of a similar gene in Caenorhabditis elegans suggests that SPY represents a class of signal transduction proteins that is present throughout the eukaryotes.

Un software FOSS para la gestión de las fotografías obtenidas por fototrampeo

La constante innovación tecnológica y al abaratamiento de los dispositivos, han hecho que el fototrampeo se convierta en una técnica ampliamente utilizada para la realización de inventarios biológicos, generando gran cantidad de registros de alto interés ecológico. El principal objetivo de esta comunicación es presentar una herramienta de software libre (FOSS), desarrollada para facilitar el manejo de datos, obtenidos mediante fototrampeo en un espacio protegido de la Comunidad Valenciana. Así, a partir de un estudio iniciado en 2009, se ha analizado la composición de la comunidad de vertebrados en el Parque Natural de la Sierra de Mariola. Para la realización de los muestreos se han empleado cámaras de infrarrojos con sensor de movimiento (Moultrie Game Spy I-60®), obteniendo un total de 29,941 contactos que han servido para determinar la abundancia de las especies muestreadas en las 63 cuadrículas (2*2 Km) establecidas en la zona de estudio. La aplicación FOSS, denominada “Camera Trap Manager”, se ha diseñado para agilizar el tratamiento de esta información y funciona sobre la plataforma .NET, pudiendo ser utilizada en varios sistemas operativos. Además, este software dispone de un interfaz de usuario muy intuitivo, capacidad de extraer automáticamente distintos tipos de metadatos (fecha, hora, fase lunar, localización, temperatura, presión atmosférica, etc), basados en el estándar Exif; disponibilidad de listas taxonómicas para permitir a los usuarios etiquetar fácilmente las imágenes con las identidades de las especies, capacidad de análisis y creación de informes. El uso de esta aplicación ha permitido simplificar el procesado, análisis y edición de datos, consiguiendo una significativa reducción de los costes económicos y temporales. Mientras que en el procesado manual se alcanzaba una media de 55 fotografías/hora, el uso de esta herramienta permite procesar más de 1,000 fotografías/hora, obteniendo una mayor cantidad de información. Finalmente, se pone de manifiesto la gran utilidad de esta aplicación, que puede ser adaptada a las necesidades de otros proyectos de gestión de fauna, haciendo su uso extensivo no solamente a los profesionales del sector, sino a todos aquellos que requieran procesar gran cantidad de imágenes de fototrampeo.

Commonplace book of Jonathan Belcher, 1725-1727

This leather-bound volume contains excerpts copied by Jonathan Belcher from books he read while he was a student at Harvard. The excerpts come from a variety of sources including periodicals and contemporary publications. The inside cover has Belcher's bookplate with the motto, "Sustine. Abstine." The back cover has some additional personal information including reference to French lessons with "Mr Law Merciers," and notes of the dates when he began certain books/essays.

The complete works of Samuel Rowlands, 1598-1628, now first collected ...

v.3. Heavens glory, seeke it, &c., 1628. The famous history of Gvy erle of Warwicke, 1682. Miscellaneous poems. Notes, by Sidney J.H. Herrtage. Glossary, by Sidney J.H. Herrtage.

Novels.

Each volume has special title-page.

Works.

v. 1. Afloat and ashore.--v. 2. The bravo.--v.3. The chainbearer.--v. 4. The crater.--v.5. The deerslayer.--v. 6. The headsman.--v. 7. The Heidenmaur.--v. 8. Home as found.--v. 9. Homeward bound.--v. 10. Jack Tier.--v. 11. Lionel Lincoln.--v. 12. The Monikins.--v. 13. Mercedes of Castile.--v. 14 Miles Wallingford.--v. 15. Last of the Mohicans.--v. 16. Oak openings.--v. 17. The Pathfinder.--v. 18. The Pilot.--v. 19. The Pioneers.--v. 20. The Prairie.--v. 21. Precaution.--v. 22. The Red rover.--v. 23. The Redskins.--v. 24. Satans toe.--v. 25. The sea lions.--v. 26. The spy.--v.27. The two admirals.--v. 28. The water-witch.--v. 29. Ways of the hour.--v. 30. Wing-and wing.--v. 31. Wept of wish-ton-wish.--v. 32. Wyandotte.

The mysteries of government, or favoritism unveiled, a New-Year's present to the people of South Carolina.

"Printed somewhere and published everywhere."

An oration, pronounced on the thirtysecond [sic] anniversary of American independence, at Barre, in the county of Worcester, July 4, 1808: /

"After the oration was delivered, the Hon. Salem Town was called to the chair, and the following resolutions ... adopted by the unanimous vote of the whole, consisting of more than six hundred Federal Republicans, from various parts of the county. Among whom were our representatives in Congress from both districts, and many other official characters."--Appendix, p. [27].

European War pamphlets

A collection of miscellaneous pamphlets on World War I.