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News - 11/13/2014 - 2:48pm

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Cytoskeletal molecular motors move uni-directionally along their tracks. This poses multiple problems: How do they get to the start of the track? Once there, how do they stay there to capture cargo? A candidate for retaining the microtubule-based motor dynein at the start of its track (microtubule plus ends) is a ubiquitous regulator called Lis1. The Reck-Peterson Lab, in collaboration with the lab of Andres Leschziner (Harvard) has been seeking to answer the second question. Previously they showed that Lis1, a gene mutated in the rare neurodevelopmental disease lissencephaly, had the functional properties necessary to stall dynein on microtubules: when bound to Lis1, dynein keeps going through the ATP hydrolysis cycle that normally powers the motor yet, rather than walking, it remains tightly bound to its track. This led to the proposal that Lis1 was working as a “clutch”, uncoupling dynein’s engine (the ATP hydrolysis cycle) from its wheels (the cycle of microtubule binding and release). In the latest issue of eLife, the Reck-Peterson and Leschziner groups now shed light on the molecular mechanism of Lis1’s affect on dynein. They reported three distinct 3D cryo-electron microscopy structures of the dynein/Lis1 complex, representing the first 3D structures of dynein bound to any of its regulators. Their structures revealed that Lis1 binding to dynein causes a repositioning of dynein’s main mechanical element, the “linker”. Based on these structures they hypothesized that Lis1 physically blocks an interaction between dynein’s linker and AAA+ motor ring, known to be required for dynein to release from microtubules. Förster resonance energy transfer (FRET), single-molecule and in vivo assays supported this hypothesis. Strikingly, shortening dynein’s linker to a point where it could physically bypass Lis1 made dynein Lis1-insensitive, showing that Lis1 keeps dynein in a persistent microtubule-bound state by directly blocking progression of its mechanochemical cycle.

For more details, click here.

News - 10/02/2014 - 8:50am

The Bjorkman-Strominger-Wiley Prize was established this year by Harvard University's Department of Molecular & Cellular Biology to encourage inter-lab cooperation, in honor of Pamela Bjorkman and Jack Strominger and the late Don Wiley. While all three were working at Harvard in the 1980s, they collaborated on research into the MHC protein’s crystal structure and antigen presentation. The first winners of this award are Samara Reck-Peterson and Andres Leschziner for their labs’ work on the motor protein dynein. According to MCB Chair Alex Schier, “the two labs combined their expertise in structural biology, biophysics, biochemistry and cell biology to be much more than the sum of their parts.”

News - 10/02/2014 - 8:46am

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PINK1 and PARKIN – two proteins mutated in early onset Parkinson’s Disease - are known to function in a signaling cascade that leads to ubiquitylation of mitochondrial outer membrane proteins on damaged mitochondria, but the precise mechanism through by which PINK1 activates PARKIN ubiquitin ligase activity and retention on the mitochondrial membrane is poorly understood. In the most recent issue of Molecular Cell, Alban Ordureau in the Harper Lab used quantitative proteomics and a technique called ubiquitin AQUA to examine the kinetics and specificity of PINK1 and PARKIN-dependent ubiquitin chain synthesis on damaged mitochondria in vivo. Through mechanistic and biochemical analysis, as well as live-cell imaging, the authors define multiple steps in the process, revealing a feed-forward mechanism for PARKIN activation and retention on mitochondria. PINK1 phosphorylation of S65 in the UBL of PARKIN leads to activation of its ubiquitin ligase activity by 2400-fold, which in turn promotes the initial synthesis of K6, K11, K48, and K63 ubiquitin chains on mitochondria by PARKIN. Ubiquitin units within newly synthesized ubiquitin chains are then phosphorylated by PINK1 on S65, the residue homologous to S65 in the UBL of PARKIN. This, in turn, serves as a binding site (Kd = 17 nM) for activated PARKIN, leading to retention of PARKIN on poly-ubiquitinated mitochondria, which could support both further ubiquitin chain synthesis and recruitment of proteins to promote mitophagy.  Our data reveal a feed-forward mechanism that explains how PINK1 phosphorylation of both PARKIN and poly-UB chains synthesized by PARKIN drives a program of PARKIN recruitment and mitochondrial ubiquitylation in response to mitochondrial damage. This work also provides a framework for quantitative analysis of phosphorylation and ubiquitin dependent signaling systems in vitro and in vivo.

News - 09/02/2014 - 10:27am

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Phosphorylation of proteins on tyrosine is particularly important in the control of cell proliferation and differentiation, and drives many of the changes seen in cancer cells. Protein tyrosine phosphorylation has previously been thought to occur only inside cells, where it can control changes in cell structure, movement, and gene expression. In a recent article in Cell, the Whitman Lab found a new kind of tyrosine kinase, VLK, that is secreted into the extracellular environment. VLK phosphorylates a broad range of extracellular proteins, including key regulators of tumor invasion and metastasis, blood clotting, and inflammation, pointing to a new mechanism for the control of protein function outside of cells.

News - 08/22/2014 - 9:56am

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Hummingbirds are avid nectar drinkers, and their ability to perceive sugars enabled their extensive radiation in a new ecological niche. In a recent article in Science, Maude Baldwin, a visiting scientist in the Liberles Lab, uncovered a noncanonical mechanism for sweet taste detection that evolved in hummingbirds since divergence from swifts, their insect-eating relatives. Phylogenetic analysis indicates that an ancestor of birds- likely within Dinosauria- lost an essential subunit of the only known vertebrate sweet receptor, raising questions of how specialized nectar-feeders sense sugars. Receptor expression studies revealed that the ancestral umami receptor (T1R1-T1R3 heterodimer) was repurposed in hummingbirds to function as a carbohydrate receptor. Furthermore, the molecular recognition properties of T1R1-T1R3 guided taste behavior in captive and wild hummingbirds. These studies illustrate how changing the ligand recognition properties of a single sensory receptor can facilitate the evolution of new species. 

For more details, see also http://hms.harvard.edu/news/sweet-feat.

News - 07/28/2014 - 10:51am

J. Wade Harper

Wade Harper has been named Chair of the Department of Cell Biology at Harvard Medical School, effective Nov. 3, 2014. He succeeds Joan Brugge who will be stepping down to co-direct the Harvard Ludwig Center.

Wade Harper is the Bert and Natalie Vallee Professor of Molecular Pathology. He received his Ph.D. in chemistry from the Georgia Institute of Technology in 1984. In 1988, Harper joined the Department of Biochemistry at Baylor College of Medicine and was recruited to the HMS Department of Pathology in 2003. He then moved to the Department of Cell Biology in 2011.

Joan Brugge, the Louise Foote Pfeiffer Professor of Cell Biology, has been a member of the HMS faculty since 1997 and chaired the Department of Cell Biology for the past 10 years.

For more details, please see HMS news coverage.

News - Jul 23, 2014

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Microtubules are fundamental for the spatial organization and motility of neurons and other cells.  In a recent article in Cell, the Flanagan Lab identified a novel function for the microtubule plus-end protein Adenomatous polyposis coli (APC), a scaffold protein known to be important in biology and disease. They found that APC is an RNA-binding protein, and identified an mRNA interactome, which was highly enriched for APC-related functions, including microtubule organization, cell motility, cancer and neurologic disease. Among the targets were tubulin mRNAs, and further studies showed that regulation of β2B-tubulin mRNA is critical for dynamic microtubule extension in axons, and for cortical neuron migration in vivo.  These results lead to a novel protein-synthesis-based model for microtubule assembly, and identify APC as a platform linking protein and mRNA networks in normal and disease states.

News - Jun 18, 2014

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A systematic quantitative analysis of temporal changes in host and viral proteins throughout the course of a productive infection could provide dynamic insights into virus-host interaction. In a recent article in Cell, the Gygi Lab describes a novel proteomic technique, ‘quantitative temporal viromics’ (QTV), employing multiplexed tandem mass tag-based mass spectrometry. They apply this technology to human cytomegalovirus (HCMV), not only an important pathogen but a paradigm of viral immune evasion. QTV detailed how HCMV orchestrates the expression of >8,000 cellular proteins, including 1,200 cell surface proteins, to manipulate signaling pathways and counter intrinsic, innate, and adaptive immune defenses. QTV predicted novel natural killer and T-cell ligands, as well as 29 viral proteins present at the cell surface--potential new therapeutic targets. Temporal profiles of >80% of HCMV canonical genes and 14 non-canonical HCMV ORFs were defined. QTV is a powerful, novel method that can yield important insights into viral infection, and is applicable to any virus with a robust in vitro model.  This work was in collaboration with the Wilkinson Lab (Cardiff University, UK) and Lehner Lab (Cambridge University, UK).

Figure: schematic of QTV strategy, as applied to HCMV.

News - Jun 10, 2014

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Cytoplasmic dynein is the major motor protein that transports cargoes toward the minus end of microtubules. In most cell types, this corresponds to movement toward the cell interior. However, if dynein only moves to the minus end of microtubules, a problem arises: how would dynein initially reach the plus end of the microtubule and the outskirts of the cell, where it collects cargoes? In a recent eLIFE article, the Reck-Peterson Lab reveals that a group of three proteins can solve this problem by transporting dynein to the plus end of the microtubule. The proteins comprise a plus-end-directed kinesin motor, and two additional proteins that connect dynein to the kinesin. Imaging the transport process shows that the dynein motor is not a passive passenger: it is able to resist against the kinesin. However, an additional microtubule-associated protein can help the kinesin motor to win this “tug of war”, and so the protein complex—including the dynein motor—moves toward the plus end of the microtubule.

Figure: Kymograph showing a fluorescently labeled dynein construct (green) moving toward the plus end of the microtubule.

News - Jun 10, 2014

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In a recent article in Nature, the Pellman Lab reports that increased numbers of centrosomes can mimic and enhance the effects of a breast cancer oncogene. Centrosome amplification, a common cytoskeletal aberration in cancer cells, has long been recognized as a feature of human tumors, but its role in tumorigenesis has been unclear.  This amplification is poorly tolerated in non-transformed cells; extra centrosomes are spontaneously lost in the absence of selection. Thus, the high frequency of centrosome amplification, particularly in more aggressive tumors, presents something of a paradox and raised the possibility that extra centrosomes could confer advantageous characteristics that promote tumor progression.

Using a 3D model system and other approaches to culture human mammary epithelial cells, the Pellman Lab found that centrosome amplification triggers cell invasion. This invasive behavior is similar to that induced by overexpression of the breast cancer oncogene ERBB2 and enhances invasiveness triggered by high-level expression of ERBB2. Their data indicate that, through increased nucleation of microtubules from centrosomes, centrosome amplification increases the activity of the small GTPase, Rac1, disrupting normal cell-cell adhesion and promoting invasion. Activation of Rac is a well-established feature of cancer cells and is thought to be critical for cancer cell invasion and metastasis.  These findings demonstrate that centrosome amplification, a structural alteration of the cytoskeleton, can promote features of malignant transformation.

 

This work was in collaboration with the Thery Lab (Hospital Saint Louis), Brugge Lab (HMS, Cell Biology), and Polyak Lab (HMS & DFCI).  

The image illustrates the ability of centrosome amplification to promote invasive behavior of mammary epithelial cells.  Top image: normal, early stage  organoids in 3-D cultures; bottom image: organoids from cells with centrosome amplification. Laminin V is in green, actin is in red, and DNA is in blue.

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