For our Journal Club on June 3, Daphné Dambournet presented the following paper:
Divergent modes for cargo-mediated control of clathrin-coated pit dynamics. Soohoo AL, Puthenveedu MA. Mol Biol Cell. 2013 Jun;24(11):1725-34. PMID: 23536704.
Model for divergent modes of cargo-mediated control of endocytosis by signaling receptors.
For our Journal Club on April 29, Sun Hae Hong presented the following paper:
The molecular basis for the endocytosis of small R-SNAREs by the clathrin adaptor CALM. Miller SE, Sahlender DA, Graham SC, Höning S, Robinson MS, Peden AA, Owen DJ. Cell. 2011 Nov 23;147(5):1118-31. PMID: 22118466
Schematic representation of the model of VAMP8 trafficking from the plasma membrane.
For our new Microtubules and Mitosis Journal Club on April 25, members of the Drubin/Barnes Lab, Eva Nogales Lab, and Ahmet Yildiz Lab discussed the following paper, selected by Nate Krefman:
Estimating the microtubule GTP cap size in vivo. Seetapun D, Castle BT, McIntyre AJ, Tran PT, Odde DJ. Curr Biol. 2012 Sep 25;22(18):1681-7. PMID: 22902755
Example EB1 comet from an EB1-EGFP LLCPK1 cell and the corresponding fluorescence intensity (FI) linescan, blue line.
Genetic Engineering & Biotechnology News is out with a Feature Article this week including some comments from David Drubin about targeted genome modification in mammalian cells for fluorescence microscopy studies.
Lighting Up Live Cells with Fluorescence. Christine Herman. GEN. Sep 1, 2012 (Vol. 15, No. 32)
“The difference between taking snapshots of the process and watching a movie is just night and day,” says David Drubin, Ph.D., professor of cell and developmental biology at the University of California, Berkeley, whose lab uses fluorescence to understand the intricate details underlying clathrin-mediated endocytosis.
Researchers in David Drubin’s lab at the University of California, Berkeley genetically engineered a human cell line to express endogenous levels of RFP-tagged clathrin light chain A (red) and GFP-tagged dynamin 2 (green) for studying clathrin-mediated endocytosis. The above 3D kymograph of the cell surface, with the time dimension in the z-axis, shows the full lifetime of hundreds of clathrin patches on the membrane, which terminate upon recruitment of dynamin. [Aaron T. Cheng]
For our next Journal Club, our summer undergraduate researcher Josh Johnson will present the following paper:
Aurora B dynamics at centromeres create a diffusion-based phosphorylation gradient. Wang E, Ballister ER, Lampson MA. J Cell Biol. 2011 Aug 22;194(4):539-49. PMID: 21844210.
Centromeres (CB-mCherry) and chromosomes (YFP emission) for a single time point. (bottom) Color-coded YFP/CFP emission ratio at different time points. The timestamp (minutes and seconds) is relative to ZM washout at t = 0. Bar, 5 µm.
The May 2012 edition of Biowire, a publication of Sigma-Aldrich, includes an interview with David Drubin about the projects in our lab looking at clathrin-mediated endocytosis (CME) in mammalian cells using zing finger nuclease (ZFN) technology to undertake targeted genome modification. Traditionally, CME has been studied in cells in which fluorescently-tagged components of endocytic machinery are overexpressed using exogenous constructs. Data obtained in many labs using these methods suggested that CME was highly variable. Using ZFN technology, in collaboration with Sangamo Biosciences, our lab recently showed that CME is robust and efficient in mammalian cells. The new results highlight the technical advantages of tagging genes at their endogenous loci, an approach that has been historically limited to genetically tractable organisms, such as the Drubin/Barnes Lab favorite Saccharomyces cerevisiae (budding yeast). Emerging technologies, such as ZFNs and TALENs, however, are now making this sort of precise genomic manipulation possible in animal cells, including human cells, giving us new and powerful ways of studying cellular biology.
Cellular processes should be studied as close to their natural states as possible. I suspect that, as we see more uses of zinc finger nucleases [for tagging endogenous genes], people will find that they have been inadvertently perturbing the processes that they have been studying.
David Drubin (Biowire, May 2012)
Prof. David Drubin, for Biowire