The C-terminal tails of heterotrimeric kinesin-2 motor subunits directly bind to α-tubulin1: Possible implications for cilia-specific tubulin entry.

The assembly of microtubule-based cytoskeleton propels the cilia and flagella growth. Previous studies have indicated that the kinesin-2 family motors transport tubulin into the cilia through intraflagellar transport. Here, we report a direct interaction between the C-terminal tail fragments of heterotrimeric kinesin-2 and α-tubulin1 isoforms in vitro. Blot overlay screen, affinity purification from tissue extracts, cosedimentation with subtilisin-treated microtubule and LC-ESI-MS/MS characterization of the tail-fragment-associated tubulin identified an association between the tail domains and α-tubulin1A/D isotype. The interaction was confirmed by Forster's resonance energy transfer assay in tissue-cultured cells. The overexpression of the recombinant tails in NIH3T3 cells affected the primary cilia growth, which was rescued by coexpression of a α-tubulin1 transgene. Furthermore, fluorescent recovery after photobleach analysis in the olfactory cilia of Drosophila indicated that tubulin is transported in a non-particulate form requiring kinesin-2. These results provide additional new insight into the mechanisms underlying selective tubulin isoform enrichment in the cilia.

Heterotrimeric kinesin-2, together with kinesin-1, steers vesicular acetylcholinesterase movements toward the synapse.

Acetylcholinesterase (AChE), which is implicated in the pathophysiology of neurological disorders, is distributed along the axon and enriched at the presynaptic basal lamina. It hydrolyses the neurotransmitter acetylcholine, which inhibits synaptic transmission. Aberrant AChE activity and ectopic axonal accumulation of the enzyme are associated with neurodegenerative disorders, such as Alzheimer's disease. The molecular mechanism that underlies AChE transport is still unclear. Here, we show that expression of Drosophila AChE tagged with photoactivatable green fluorescent protein and m-Cherry (GPAC) in cholinergic neurons compensates for the RNA interference-mediated knockdown of endogenous AChE activity. GPAC-AChE, which is enriched in the neuropil region of the brain, moves in the apparently vesicular form in axons with an anterograde bias in Drosophila larvae. Two anterograde motors, kinesin-1 and -2, propel distinct aspects of GPAC-AChE movements. Total loss of kinesin-2 reduces the density of anterograde traffic and increases bidirectional movements of GPAC-AChE vesicles without altering their speed. A partial loss of kinesin-1 reduces both the density and speed of anterograde GPAC-AChE traffic and enhances the pool of stationary vesicles. Together, these results suggest that combining activity of a relatively weak kinesin-2 with that of a stronger kinesin-1 motor could steer AChE-containing vesicles toward synapse, and provides a molecular basis for the observed subcellular distribution of the enzyme.-Kulkarni, A., Khan, Y., Ray, K. Heterotrimeric kinesin-2, together with kinesin-1, steers vesicular acetylcholinesterase movements toward the synapse.

Developing a faculty support program for fostering enriching undergraduate laboratory experiences under limited resource conditions.

Meaningful pedagogical reform requires good faculty training and support programs. Such support is particularly valuable when colleges and universities are trying to bring research and inquiry into the laboratory curricula under resource-limited conditions. In this situation, it may help to extend the scope of the faculty support program to include training for practicing experimental techniques, sustainable networking opportunities, and a space to learn about pedagogical reforms. From this perspective, we share our experience about building a faculty development program for public college teachers who teach undergraduate biology in India. Though we designed the program for low-resource settings, the experiments curated could very well represent core biological concepts typically identified by the international community. The activities and overall design of the program can be useful for initiating pedagogical reform in any college/university where the traditional approach to biology laboratory instruction predominates, and high-end research is not easy to access.

Preparation and Testing of Food Freshness Indicators: an Application-Oriented Learning Module Integrating Basic Concepts of Microbiology and Chemistry Laboratory.

We describe a novel 4-day food microbiology laboratory learning module for a first-year, introductory undergraduate course. In the module, the students test the suitability of four different pH indicator dyes as freshness indicators for dairy products. The concepts of serial dilutions, microbial growth, microbial metabolism, pH as well as pKa, and basic microbial laboratory practices are a part of the designed activity. It is a relatively inexpensive module and can be executed with little infrastructural support. It can be delivered as a stand-alone structured inquiry. The associated variables and applications indicate that the activity can perhaps be developed into a more elaborate course-based undergraduate research experience, or CURE.

A module integrating conventional teaching and student-centered approach for critical reading of scientific literature.

One of the well-accepted ways of introducing undergraduates to the method of scientific inquiry is to introduce them to primary literature. A matter of concern is that undergraduate students of Indian universities are generally not trained to read primary literature. We combined traditional teaching and student-centered approach in a 3-day module for introducing first-year students to critical reading of primary literature. This article describes the design of the module, the method of assessment of student performance, the learning outcomes, and student feedback. We received positive feedback from students and observed significant improvement in their understanding of the rationale and results of the research paper under consideration. The module may be viewed as an example of a simple introductory activity to enthuse and initiate college students to scientific thinking and research paper reading. © 2019 International Union of Biochemistry and Molecular Biology, 47(5):581-588, 2019.