ewinter's blog

Receptor tyrosine kinases (RTKs) are a class of membrane-spanning proteins that dimerize when a dimer signaling molecule binds the extracellular receptor domain. When two RTKs dimerize, they autophosphorylation on multiple tyrosine residues. The adaptor protein GRB2 binds to a phosphorylated tyrosine via its SH2 domain. Ras is anchored to the lipid bilayer and is a kinase for many downstream growth pathways. SOS is a Ras-GEF, so upon binding Ras, changes its conformation to have less affinity for GDP and more for GTP so it can get a phosphate group and phosphorylate things such as the Map kinases. The RTK/Ras/MapK pathway has been implicated at multiple points to be overactive in cancer. Drugs such as dacomitinib work to irreversibly bind cystines in the ATP binding kinase domain of the RTK. However, secondary driver mutations often arise in downstream targets such as Ras, rendering treatment of the RTK useless in treating cancer.

Henry Molaison, commonly known as “Patient H.M.” was a man who underwent a bilateral medial temporal lobectomy to treat violent seizures. After this surgery, he had severe anterograde amnesia and some retrograde amnesia. The medial temporal lobe became known as the hippocampus, and we learned from Patient H.M. that is is important in encoding declarative memories, but not procedural memories. The hippocampus is also important for storing spatial memories, and place cells are known to fire both when rats are at a specific location, and during sleep so these spatial memories can be consolidated.

Long term potentiation is the form of learning at the synaptic level, by strengthening of a specific synapse. Induction of these changes results from repeated, high frequency stimulation of a synapse, also known as tetanus. The stimulated AMPA receptors open with glutamate binding, and allow Na2+ into the cell. When enough of these are opened, the voltage-gated NMDA receptors become unblocked by Mg2+ and allow Ca2+ in. NMDA receptors need both glutamate and depolarization to open. This elicits a positive feedback loop in which retrograde gaseous neurotransmitters like nitric oxide are produced to stimulate glutamate release in the presynaptic cell. Late phase long-term potentiation is characterized by structural changes such as fatter synapses, more dendritic spines, more terminal buttons, and a larger number of synapses.

How animals learn to communicate is a very interesting topic. For example, it has been shown that songbirds raised in isolation produce sounds that are consistent with their species. The Wernicke-Geschwind model of language says that language is a result of an interconnected network of components. Language processing is left lateralized in the brain. There are two types of aphasia: Broca’s aphasia and Wernicke’s aphasia. Broca’s aphasia is due to damage to the left frontal lobe, and is characterized by difficulty in language production but not in comprehension. Wernicke’s aphasia is caused by damage to the left superior temporal gyrus, and is characterized by difficulty with language comprehension but not production. The arcuate fasciculus is a bundle of fibers connecting Wernicke’s area and Broca’s area.

Signal transduction, in a sensory processing sense, is the conversion of energy into a neural signal. It occurs in receptor cells located in sensory organs such as the ears, eyes, and hands. Receptor cells are responsive to certain types of energy, but not others. In the cochlea (inner ear) hair cells located in the basilar membrane have stereocilia, which are hair-like structures that touch the tectorial membrane. Sound vibration causes hair displacement and opens mechanically gated ion channels, which causes the cells to depolarize and release neurotransmitters. These cells do not fire action potentials. There are four different types of touch receptors: pain, touch, vibration, and stretch. These can be found subcutaneously all around the body. Each touch receptor type has a distinct pathway to the brain. The visual system detects both brightness and contrast. Photoreceptors perform signal transduction. There are two types of photoreceptors: scotopic (rods), which work in dim light, and photopic (cones), which govern vision of colors. The visual pathway crosses sides at the optic chiasm, so the right visual field is processed in the left occipital lobe, and vice versa.

There are two types of processing: top-down and bottom-up control. Top-down processing is when the brain communicates to the organ based on previous experience. Bottom-up processing is the opposite, when information from the environment is relayed to the brain. The concept of attention is preferential processing of a subset of information from the environment. Endogenous attention is voluntary, top-down control that can be sustained over long periods of time. The main brain region involved is the frontoparietal networks. Exogenous attention is reflexive, bottom-up processing which ir rapid but quickly fades. It primarily uses modality-specific brain areas, such as the primary visual cortex or auditory cortex.

Signal transduction, in a sensory processing sense, is the conversion of energy into a neural signal. It occurs in receptor cells located in sensory organs. These sensory organs include the eyes and the ears. In the cochlea (inner ear) hair cells located in the basilar membrane have stereocilia, which are hair-like structures that touch the tectorial membrane. Sound vibration causes hair displacement and opens mechanically gated ion channels, which causes the cells to depolarize and release neurotransmitters. These cells do not fire action potentials.

Using the Primer3 software, primers were created to amplify the region of the NaN1793 mutation. These primers were chosen with respect to the fact that Sanger sequencing can only sequence about 1000 b.p. maximum, and the mutation should not be too close to either primer since the beginning and end of sequencing data is usually less accurate than the middle. The expected size of the amplicon is 874 b.p. In order to confirm that DNA was extracted from the plants and to decide which samples to take for sequencing, gel electrophoresis was run. Two gels were run. One gel contains PCR products in a ¼ dilution in T10E1 (TE) buffer while the other contains PCR products in a ¼ dilution with H2O as a control. The T10E1 diluted DNA was preferentially taken for sequencing because it protects DNA from degradation, so that DNA should be of better quality than the H2O diluted DNA. Because there was no band in the Mutant 1 lane of the TE gel, Mutant 1 was taken from the H2O gel. Mutant 2, Mutant 6, and Mutant 7 were all taken from the TE gel. The DNA was extracted and purified from the gel and was nanodropped to confirm presence of DNA and assess purity. In order to perform Sanger sequencing, a primer is needed. The forward primer is 428 b.p. from the mutation site, while the reverse primer is 447 b.p. from the mutation site. The forward primer was used for sequencing because it is closer to the mutation site.

Toluidine blue (T-blue) stains polysaccharides and phloroglucinol HCL (Ph-HCL) stains lignin. In the NaN1793 mutants, both the Ph-HCL and T-blue stains appear a much lighter color at the distal cortex as opposed to the proximal cortex (Fig. 7B, 7D). In the wild - type plants, this contrast is not observed to as great of an extent (Fig. 7). The NaN1793 mutation is predicted to be a nonsense mutation, resulting in a truncated, nonfunctional protein. This phenotypic difference suggests that Bradi1g25180 may be involved in formation of cell walls of the distal cortex.

Using the Primer3 software, primers were created to amplify the region of the NaN1793 mutation. These primers were chosen with respect to the fact that Sanger sequencing can only sequence about 1000 b.p maximum, and the mutation should not be too close to either primer since the beginning and end of sequencing data is usually less accurate than the middle. The expected size of the amplicon is 874 b.p. In order to confirm that DNA was extracted from the plants and to decide which samples to take for sequencing, gel electrophoresis was run. Two gels were run. One gel contains PCR products in a ¼ dilution in T10E1 (TE) buffer while the other contains PCR products in a ¼ dilution with H2O as a control. T10E1 buffer protects nucleic acids from degradation.