Nanotechnology in Single Cell Biology & Cellular Processes in Single Cell: Cell Cycle

It’s been raining nonstop since I landed in Taipei – not storming but a constant pitter patterning of rain drops. I don’t mind it. It’s actually kind of nice and soothing. Especially with the poster session taking place in a covered, outdoor courtyard, the rain adds an almost zen element, contributing to a very relaxing environment to discuss Science!

 

Going back to this morning, I got lost trying to find the lecture hall on campus. Luckily, a very nice campus security guard took pity on me and my poor attempt at speaking Mandarin and was able to point me in the right direction. I made it just in time for the start of the morning session, which kick started with the Advanced Microscopy for Single Cells and was followed by Single-Cell Mechanobiology I (Michael will be covering these sessions).

 

Lunch followed, and I couldn’t help but overflow my plate with food to the point where it was almost embarrassing (the key word is almost). This boost of energy sustained me through the poster session (take that jetlag!). The Nanotechnology in Single Cell Biology session followed immediately afterwards, where Dr. Bianxiao Cui started by posing the key biological question of “How do cells sense surface topography?”. To help answer this overarching question, the Cui group etches tunable nanopillar features into glass and has discovered that certain proteins preferentially accumulate on the nanopillars. One important class of proteins are those that contribute to clathirin-mediated endocytosis (i.e. FCHo1, TfnR, Epsin1, Amphiphysin1, CLTA, DNM2, and AP2). Moreover, when cells are cultured on nanobar features that possess high curvature at the end and low curvature along its length, these endocytic-related proteins display distinct curvature preferences by accumulating at the ends of the nanobars. The resulting curvature induced by the features was found to also affect endocytosis kinetics. To read more about this work, please refer to Dr. Cui’s recently published paper. Another important class of proteins that the Cui group discovered to be sensitive to curvature were those responsible for forming branched actin, such as FBP17, N-WASP, and Cortectin.

 

Dr. Haw Yang then proceeded to share his key biological question of “How do molecular interactions give rise to larger-scale processes?” or, more specifically, “How do rare events frame the mechanisms and kinetics of biophysical interactions?”. In order to address these types of questions, Dr. Yang presented a multi-resolution imaging approach that combines two-photon laser scanning microscopy and 3D single-particle tracking to investigate nano-bio interactions in living single cells. One application that Dr. Yang demonstrated is the ability to provide mechanistic insights for cellular trafficking of nanoparticles. For example, his group discovered that TAT-coated nanoparticles move on the membrane surface rather than inside of the endosome in addition to there being a faster-than-expected slowdown of these nanoparticles when approaching the cell membrane.

 

Scott Thourson ended the nanotechnology session by presenting tunable, conductive PEDOT:PSS polymer microwires. He started with a demonstration that these microwires were capable of stimulating a single heart cell, which could be visualized by the “beating” or contraction of the cell. Thourson then proceeded to motivate the use of these polymer microwires in combination with the patch clamp method in order to obtain more quantitative results about membrane potential. Since these wires are comprised of a soft conductor (Young’s modulus around 2 Pa, similar to brain tissue) with high charge density, they are ideal for use in single neuron stimulation, which Thourson highlighted is sufficient to affect behavior. Moreover, these polymer wires can be grown with tunable properties that can influence stimulation parameters, making them a promising interfacial material to interact electrically with single cells.

 

After a brief coffee break, the afternoon continued with the Cellular Processes in Single Cell: Cell Cycle session. Dr. Paul Wiggins kick-started the session discussing the “wrestling match” that occurs between replication and transcription and his group’s interest in understanding how frequent these replication conflicts are as well as what are the structural consequences of such conflicts. Using Single-Molecule Fluorescence Microscopy, the Wiggins group is able to visualize the replication process in single cells and characterize the replication complexes with single-molecule resolution, revealing that 1) replication is inherently discontinuous with pervasive disassembly/assembly and 2) transcription-induced conflicts are a key contributing factor for replisome disassembly. Furthermore, it appears that replication conflicts are actually quite frequent (5 conflicts per cell cycle). For more information regarding this work, please refer to their recently published work.

 

Dr. Sheng-Hong Chen started his talk by motivating the importance of p53 and MDMX in cancer, stating that the tumor supressor p53 is mutated in over 50% of cancer patients and that the oncogene, MDMX, is often overexpressed. Therefore, current therapeutic approaches to combating cancer often involve the dual approach of activating p53 while inhibiting MDMX. The success of this approach, however, hinges on the timing of the activation/inactivation. In order to obtain a better understanding of the optimal temporal parameters, Dr. Chen began by dissecting the dynamics of p53 in single cells. Upon exposure to gamma irradiation, Dr. Chen described an oscillatory behavior of p53 that followed with cell-cycle arrest. On the other hand, with UV irradiation, there was a single pulse of p53 that triggered apoptosis. The Chen group is interested if it is possible to re-direct p53 dynamics by modulating MDMX. MDMX suppression was found to trigger biphasic p53 dynamics in single cells: 1) an initial post-mitotic pulse and 2) an oscillation phase. Furthermore, cell sensitivity to DNA damage is also different in each of the phases. In the first, MDMX works in synergy with DNA damage to cause cell death while, in the second, MDMX inhibited cell death by re-directing UV-induced apoptosis to cell cycle arrest — highlighting the key take away that timing is important!

 

Olivia J. Scheideler (University of California – Berkeley)

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