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Biomedical subjects

M R Hanson

Publications and source records attributed to M R Hanson.

At least 19 recordsLinked to original sources
PubMed2005

Enhanced protein kinase B/Akt signalling in pituitary tumours.

Pituitary tumours have previously been shown to harbour several abnormalities that cause deregulation of the cell cycle, particularly down-regulation of expression of the cyclin-dependent kinase inhibitor p27. However, it has been unclear whether these are the primary initiating events, or are secondary to other more proximate alterations in signalling pathways. In other cellular systems the Akt signalling pathway has been associated with downstream modulation of cell-cycle control. The aim of the present study was to test the hypothesis that Akt signalling is enhanced in pituitary tumours, and to see if changes in Akt expression are related to previous findings on low expression levels of the nuclear cell-cycle inhibitor p27 in pituitary tumours. We examined normal and adenomatous human pituitary tissue for mRNA and protein expression of Akt1, Akt2 and p27, and the activation of Akt, as well the phosphatase involved in the inactivation of Akt, phosphatase and tensin homologue deleted on chromosome 10 (PTEN). In pituitary adenomas Akt1 and Akt2 mRNA were found to be over-expressed compared with normal pituitary, while PTEN transcripts showed similar levels between the two tissue types. Immunohistochemical expression of phospho-Akt was found to be higher in the tumours than normal pituitaries, while the protein expression of nuclear p27 and PTEN was lower in the adenomas. However, the expression of p27 and Akt were not directly correlated. PTEN sequencing revealed no mutation in the coding region of the gene in pituitary adenomas, and thus we did not locate a cause for the increased phosphorylation of Akt. In summary, we have shown over-expression and activation of the Akt pathway in pituitary tumours, and we speculate that cell-cycle changes observed in such tumours are secondary to these more proximate alterations. Since Akt is a major downstream signalling molecule of growth factor-liganded tyrosine kinase receptors, our data are most compatible with an abnormality at this level as the primary driver of pituitary tumorigenesis.

Cell Cycle Proteins↗
PubMed2004

Stromules and the dynamic nature of plastid morphology.

Investigation of plastids via green fluorescent protein (GFP) has led to the rediscovery of tubular extensions of the plastid membrane, termed stromules, for stroma-filled tubules. These unique structures are challenging our understanding of plastid structure and function. Stromules are highly dynamic, branching and elongating across the plant cell. Recent experiments indicate that cytoplasmic microtubules and microfilaments control the shape and motility of stromules. Whether stromule formation involves plastid-specific structural systems, such as the plastid division machinery, remains open to debate. Fluorescence photobleaching experiments have revealed that GFP can traffic between plastids joined by stromules. As a result, interest has grown in whether other macromolecules can also travel through these connections. Although the function of stromules is unknown, several aspects of their biology suggest they play a role in molecular exchange between plastids and other organelles.

Arabidopsis Proteins↗
PubMed2001

Edited transcripts compete with unedited mRNAs for trans-acting editing factors in higher plant chloroplasts.

Chloroplast RNA transcripts of vascular plants undergo C to U editing at approximately 30 sites, but there is no consensus sequence that identifies a C to be edited. Both sequences closely surrounding an edited C and unidentified site-specific trans-acting factors have been shown to be important for editing. The ability of an already edited transgenic sequence to bind and thus titrate a trans-acting editing factor was evaluated for two editing sites, ndhF and rpoB site 2. The U-containing rpoB transcripts did not affect editing of the endogenous rpoB transcripts, likely because the comparable C-containing transcripts containing 27 nucleotides surrounding the edited C were only 20% edited, indicating a low affinity of a trans-factor for this length of edited sequence. Surprisingly, U-containing ndhF transgene transcripts reduced endogenous ndhF transcript editing to the same degree as a C-containing transgene transcript. This indicates that the C target of editing is not a critical recognition feature for the site-specific trans-acting factor.

Binding Sites↗
PubMed2001

A single alteration 20 nt 5' to an editing target inhibits chloroplast RNA editing in vivo.

Transcripts of typical dicot plant plastid genes undergo C-->U RNA editing at approximately 30 locations, but there is no consensus sequence surrounding the C targets of editing. The cis-acting elements required for editing of the C located at tobacco rpoB editing site II were investigated by introducing translatable chimeric minigenes containing sequence -20 to +6 surrounding the C target of editing. When the -20 to +6 sequence specified by the homologous region present in the black pine chloroplast genome was incorporated, virtually no editing of the transcripts occurred in transgenic tobacco plastids. Nucleotides that differ between the black pine and tobacco sequence were tested for their role in C-->U editing by designing chimeric genes containing one or more of these divergent nucleotides. Surprisingly, the divergent nucleotide that had the strongest negative effect on editing of the minigene transcript was located -20 nt 5' to the C target of editing. Expression of transgene transcripts carrying the 27 nt sequence did not affect the editing extent of the endogenous rpoB transcripts, even though the chimeric transcripts were much more abundant than those of the endogenous gene. In plants carrying a 93 nt rpoB editing site sequence, transgene transcripts accumulated to a level three times greater than transgene transcripts in the plants carrying the 27 nt rpoB editing sites and resulted in editing of the endogenous transcripts from 100 to 50%. Both a lower affinity of the 27 nt site for a trans-acting factor and lower abundance of the transcript could explain why expression of minigene transcripts containing the 27 nt sequence did not affect endogenous editing.

Base Sequence↗
PubMed2001

Identification of a BIBAC clone that co-segregates with the petunia restorer of fertility (Rf) gene.

Molecular markers closely linked to the Restorer of fertility (Rf) locus in petunia were sought by conducting a bulk segregant analysis. The co-segregation of markers and Rf was tested on a large BC1 population produced from two different parental lines carrying Rf. The recombination frequency between OP704 and ECCA/MACT, the two most distal markers utilized in the fine-scale mapping. was significantly different in populations derived from parents that carry different nuclear backgrounds. The fine mapping identified an amplified fragment length polymorphism (AFLP) marker that co-segregates with Rf. A petunia BIBAC library (four genome equivalents), with an average insert size of 70 kb, was constructed and screened with the linked marker. A contiguous map was constructed from three different BIBAC clones that hybridized to the marker. As a result, we have identified a 37.5-kb BIBAC clone that co-segregates with Rf.

Chromosome Mapping↗
PubMed2001

High-level expression of a synthetic red-shifted GFP coding region incorporated into transgenic chloroplasts.

We describe here a synthetic red-shifted variant of GFP that can be introduced into tobacco plastid genomes and is highly expressed in regenerated plants that appear normal and fertile. The variant contains the S65G and S72A mutations which shift the absorption maximum from the 395 nm of wild-type GFP closer to 488 nm, a wavelength emitted by a laser commonly used in confocal microscopy. In addition to enhanced fluorescence, the removal of significant absorption below 450 nm will potentially facilitate double-labelling experiments. The variant GFP encoded by the synthetic gene can be expressed at a high level, forming approximately 5% of total leaf protein.

Amino Acid Sequence↗
PubMed2001

Catechol-O-methyltransferase inhibitors in the management of Parkinson's disease.

Parkinson's disease is the most common neurodegenerative disease in which the chemical pathology is known and effective symptomatic treatment, levodopa, is available. Therapy in the initial years after initiation with dopa decarboxylase inhibitors, carbidopa or benserazide, combined with levodopa results in favorable, stable responses. However, by 5 years after the initiation of treatment, over two thirds of patients experience motor fluctuations beginning initially with a "wearing-off" effect followed by more complex fluctuations including dyskinesias and "on-off" responses. A number of strategies have been developed in an attempt to deal with these complications including changing doses and frequencies, adding agonist medications, adding or substituting controlled-release levodopa, and surgical therapies. A more recent strategy has centered on increasing the availability of intracellular levodopa and synaptic dopamine by inhibiting the peripheral and central metabolism of levodopa to 3-O-methyldopa with the use of a catechol-O-methyltransferase inhibitor. To date, two of these inhibitors, tolcapone and entacapone, are available to treat the wearing-off phase of levodopa therapy.

Benzophenones↗
PubMed2001

Unusual causes of hemifacial spasm.

Hemifacial spasm (HFS) has been defined as consisting of brief clonic jerking movements of the facial musculature, beginning in the orbicularis oculi with downward spreading to other facial muscles. HFS, perhaps the most common of the abnormal involuntary facial movements, has been classically ascribed to vascular loop compression at the root exit zone of the facial nerve. Causes other than such vascular loops are rare in the medical literature. Here we present three case studies in which the phenomenology of the HFS was atypical in onset and evolution. Using these three patients as introduction to the topic, we reviewed the literature of all cases of HFS with causes other than the vascular loop. In these three cases, HFS was caused by (1) a parotid gland tumor, (2) a cerebellopontine angle meningioma, and (3) an acoustic schwannoma. We also discuss the radiological findings as well as possible differences in the genesis of HFS and phenomenology in such cases and present recommendations on how to evaluate these patients.

Adenocarcinoma↗
PubMed2001

GFP imaging: methodology and application to investigate cellular compartmentation in plants.

The cloning of the jellyfish gfp (green fluorescent protein) gene and its alteration for expression in subcellular locations in transformed plant cells have resulted in new views of intracellular organization and dynamics. Fusions of GFP with entire proteins of known or unknown function have shown where the proteins are located and whether the proteins move from one compartment to another. GFP and variants with different spectral properties have been deliberately targeted to separate compartments to determine their size, shape, mobility, and dynamic changes during development or environmental response. Fluorescence Resonance Energy Transfer (FRET) between GFP variants can discern protein/ protein interactions. GFP has been used as a sensor to detect changes or differences in calcium, pH, voltage, metal, and enzyme activity. Photobleaching and photoactivation of GFP as well as fluorescence correlation spectroscopy can measure rates of diffusion and movement of GFP within or between compartments. This review covers past applications of these methods as well as promising developments in GFP imaging for understanding the functional organization of plant cells.

Cell Compartmentation↗
PubMed2000

Programmed cell death during pollination-induced petal senescence in petunia.

Petal senescence, one type of programmed cell death (PCD) in plants, is a genetically controlled sequence of events comprising its final developmental stage. We characterized the pollination-induced petal senescence process in Petunia inflata using a number of cell performance markers, including fresh/dry weight, protein amount, RNA amount, RNase activity, and cellular membrane leakage. Membrane disruption and DNA fragmentation with preferential oligonucleosomal cleavage, events characteristic of PCD, were found to be present in the advanced stage of petal senescence, indicating that plant and animal cell death phenomena share one of the molecular events in the execution phase. As in apoptosis in animals, both single-stranded DNase and double-stranded DNase activities are induced during petal cell death and are enhanced by Ca(2+). In contrast, the release of cytochrome c from mitochondria, one commitment step in signaling of apoptosis in animal cells, was found to be dispensable in petal cell death. Some components of the signal transduction pathway for PCD in plants are likely to differ from those in animal cells.

Apoptosis↗
PubMed2000

Plastid tubules of higher plants are tissue-specific and developmentally regulated.

Green fluorescent stroma filled tubules (stromules) emanating from the plastid surface were observed in transgenic plants containing plastid-localized green fluorescent protein (GFP). These transgenic tobacco plants were further investigated by epifluorescence and confocal laser scanning microscopy (CSLM) to identify developmental and/or cell type specific differences in the abundance and appearance of stromules and of plastids. Stromules are rarely seen on chlorophyll-containing plastids in cell types such as trichomes, guard cells or mesophyll cells of leaves. In contrast, they are abundant in tissues that contain chlorophyll-free plastids, such as petal and root. The morphology of plastids in roots and petals is highly dynamic, and plastids are often elongated and irregular. The shapes, size, and position of plastids vary in particular developmental zones of the root. Furthermore, suspension cells of tobacco exhibit stromules on virtually every plastid with two major forms of appearance. The majority of cells show a novel striking 'octopus- or millipede-like' structure with plastid bodies clustered around the nucleus and with long thin stromules of up to at least 40 (micro)m length stretching into distant areas of the cell. The remaining cells have plastid bodies distributed throughout the cell with short stromules. Photobleaching experiments indicated that GFP can flow through stromules and that the technique can be used to distinguish interconnected plastids from independent plastids.

Cell Differentiation↗
PubMed2000

Active protein transport through plastid tubules: velocity quantified by fluorescence correlation spectroscopy.

Dynamic tubular projections emanate from plastids in certain cells of vascular plants and are especially prevalent in non-photosynthetic cells. Tubules sometimes connect two or more different plastids and can extend over long distances within a cell, observations that suggest that the tubules may function in distribution of molecules within, to and from plastids. In a new application of two-photon excitation (2PE) fluorescence correlation spectroscopy (FCS), we separated diffusion of fluorescent molecules from active transport in vivo. We quantified the velocities of diffusion versus active transport of green fluorescent protein (GFP) within plastid tubules and in the cytosol in vivo. GFP moves by 3-dimensional (3-D) diffusion both in the cytosol and plastid tubules, but diffusion in tubules is about 50 times and 100 times slower than in the cytosol and an aqueous solution, respectively. Unexpectedly larger GFP units within plastid tubules exhibited active transport with a velocity of about 0.12 microm/second. Active transport might play an important role in the long-distance distribution of large numbers of molecules within the highly viscous stroma of plastid tubules.

Amino Acid Sequence↗
PubMed2000

Effective treatment of Alzheimer disease and its complications.

In Alzheimer disease, therapies to improve the core symptoms and perhaps even slow disease progression include cholinesterase inhibitors, receptor agonists, antiinflammatory drugs, and antioxidants. Neuroleptics, antiepileptics, and nondrug approaches are used to control and relieve complications such as delusions, hallucinations, paranoia, and agitated behavior. We outline a practical approach to the use of these therapies.

Alzheimer Disease↗
PubMed1998

Amlodipine for migraine prophylaxis.

Migraine is among the most common neurologic disorders encountered in clinical practice. In the general US population, the annual incidence has been calculated to be approximately 250 per 100,000 with a point prevalence of 10%. Females are affected more than males. A variety of prophylactic and abortive medications are being used for treatment and several are being studied in clinical trials. Calcium-channel blockers are frequently used prophylactic medications. We report two patients with migraine successfully treated with amlodipine (Norvasc, Pfizer, Inc), a slow calcium-channel blocker. To our knowledge, these are the first reported cases of amlodipine used in migraine prophylaxis.

Adult↗
PubMed1997

Exchange of protein molecules through connections between higher plant plastids.

Individual plastids of vascular plants have generally been considered to be discrete autonomous entities that do not directly communicate with each other. However, in transgenic plants in which the plastid stroma was labeled with green fluorescent protein (GFP), thin tubular projections emanated from individual plastids and sometimes connected to other plastids. Flow of GFP between interconnected plastids could be observed when a single plastid or an interconnecting plastid tubule was photobleached and the loss of green fluorescence by both plastids was seen. These tubules allow the exchange of molecules within an interplastid communication system, which may facilitate the coordination of plastid activities.

Amino Acid Sequence↗
PubMed1997

The green fluorescent protein as a marker to visualize plant mitochondria in vivo.

To determine how to utilize the green fluorescent protein (GFP) as a marker for subcellular localization and as a label for plant mitochondria in vivo, transgenic suspension cells and tobacco plants expressing GFP with and without a mitochondrial localization signal were generated. The first GFP form used, GFP1, is easily observable in cells with low autofluorescence, such as suspension cells or trichomes, but masked in green tissue. For the visualization of GFP in cells and tissues with high autofluorescence, such as leaf, the use of a very strong promoter (35S35SAMV), a highly expressed modified mGFP4 coding region and a brighter mutant form of GFP (S65T) was necessary. Confocal or two-photon laser scanning microscopy reveal a distinct subcellular localization of the fluorescence in cells expressing GFP or coxIVGFP. In cells expressing untargeted GFP, fluorescence accumulates in the nucleoplasm but is also distributed throughout the cytoplasm. It is excluded from vacuoles, nucleoli and from round bodies that are likely to be leucoplasts. In contrast, fluorescence is localized specifically to mitochondria in cells expressing coxIVGFP fusion protein as shown by co-localization with a mitochondrial-specific dye. This permits the direct observation of mitochondria and mitochondrial movements in living plant cells and tissues throughout plant development. Three-dimensional reconstruction of individual cells can give additional information about the distribution and numbers of mitochondria.

Cells, Cultured↗