Day 3- Single-Cell Mechanobiology

After the wonderful talk during coffee break, coming back to our last session of today!

Megan Valentine- A new model system for cellular studies of mechanobiology

Megan introduced us a new model organism that is known is our  “closest vertebrate relative”, Botryllus schlosseri (commonly known as the golden star tunicate). It is a highly dynamic organism that needs constant angiogenesis, because it has a large and transparent extracorporeal vascular network, and their vessels are constantly remodeling. What is special about them is that their vessels are inverted with respect to vertebrate, so we can have direct access to extracellular matrix via microinjection.

With the model organism Botryllus, she can directly apply physical forces and monitor the downstream responses in a living organism in real time through manipulation of the blood vessels. She found that Lysyll oxidase (LOX, responsible for crosslinking collagen) expression is stimulated by the presence of collagen, and inhibiting LOX by adding a specific inhibitor, ß-aminopropionitrile (BAPN) causes massive retraction of vessels.

This is a pretty fascinating new model system for mechanobiology studies, and this talk was ended with a nice and amusing “slurp” video (a cell swallowed by the phagocyte)!

 

Chin-lin Guo- Spontaneous Patterning of Cytoskeleton in Single Epithelial Cell Apicobasal Polarity Formation

How does mammalian cells form the specific organs? Previously, people focused on the spatial patterning and the coordination of chemical signals. Recently, ChinLin and others have found that mechanical forces also play an important role in the organization of multicellular architectures.

He has shown that long-range mechanical force enables self-assembly of epithelial tubular patterns, and the  self-organization of epithelial morphology is dependent on rigidity. Moreover, he thinks that direct cell-cell contact induces the segregation of par complex and the formation of actin belt, which is how individual cells form apicobasal polarity.

The cool part of his talk is that he uses an ECM gel to see the spontaneous single-cell partitioning of cytoskeleton on 2D platform and 3D culture. Furthermore, to differentiate between actin band and belt and the role of microtubules, he implemented the lattice light sheet microscopy (LLSM, Bi-Chang Chen), which showed the conversion of actin from band to contractile belt, and he thinks that actin band can serve as a precursor to guide cell-cell interactions.

To sum up, he thinks that single epithelial cells can form a precursor state by spontaneous partitioning of cytoskeleton to guide multicellular epithelization and apicobasal polarity formation, and there could be an intermediate state between the mesenchymal state and the epithelial state.

 

Poul Bendix- Dynamics of Filopodia: Rotation, Twisting, and Pulling

Poul is interested in the question: do filopodia rotate around its own axis? His group has surprising evidence for a new pulling mechanism originating from twisting of the actin within the filopodium. Using labeled actin, he can have 3D visualization of filopodia in different cells to find the answer.

When visualizing actin polymerization inside membrane tube, he found that filopodia exhibit buckling of their actin shaft in conjunction with pulling. In HEK cells, he found that there is twist buckling transition of filopodia that buckling releases accumulated twist, which is a strong indication of twisting of actin. Moreover, they observed retrograde flow and rotation of the actin shaft, so there could be correlation between force and actin distribution, and also correlation between coil movement and force.

They have found helical buckling and rotational behavior in the filopodial actins in various cell lines, which may facilitate the sensing and interaction of the cell with its surroundings using filopodia.

 

***

The banquet in Grand Hotel was AMAZING!! Grand hotel is one of the most famous landscapes of Taipei, with contemporary palatial architecture and delicious cuisines! After the wonderful meal, some of us went to the karaoke, and I heard it was also lots of fun!

Ivy (Howard Lab / Howard Lab facebook & Xiong Lab, Yale University)

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IMG_5640.JPG(Having a great meal at Grand Hotel, photo: Ivy)

 

 

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The Final Morning at the Mechanobiology Meeting

After the banquet, there was one last morning of scientific sessions.  It started with thanking those that made the meeting happen.  One of the organizers, Linda Kenney noted that over 30% of the speakers at the meeting were women, and encouraged others to strive to do the same when they plan meetings.   This was followed by an action packed morning session.

Xavier Trepat from IBEC, showed some very interesting studies to understand mechanisms of how cancer cells migrate out of tumors. Using tumor cells of epithelial origin (A341 cells) and from clinical samples of lung cancers that these cells could indeed bind using cell-cell adhesions mediated by E cadherin and N cadherin. To demonstrate that these two cadherins can form heterotypic adhesions they presented different cadherin (E,N,P) on magnetic beads to the cells. A magnetic field was applied for a minute and assayed how the percentage of beads bound to the cell. Here they saw that N-cad- Ecad adhesions were formed. These adhesions between cancer cells and CAFs(cancer activated fibroblasts) actually assist migration where the CAFs act as the leader cell and pull the cancer cells along their path of migration. Using traction force microscopy, they also observed that the leader cell actually pulls onto the CAFs. In some interesting preliminary experiments, the Trepat group also shows that when cancer cells associate with CAF, they lead to the polarization and subsequent migration of CAF by a yet unknown mechanism.

This was followed by a somewhat related talk by Kevin Chalut from the University of Cambridge. In his lab, they have developed a different method to make hydrogels of different rigidities in a more reliable manner. Changing only the stiffness of hydrogels they show that OPC proliferation relies on soft matrices. There was a nice study on comparing pre- and post- implantation tissue with stem cells grown on soft matrices. On comparison of gene expression profile, they found that pre-implantation embryos were comparable to soft gels whereas post-implantation embryos were comparable to stem cells grown on stiffer gels. Further they went on to show that in the brain, regeneration occurs better when the matrix is softer. Using decellularized tissues, they showed that ageing matrices are more stiff and support proliferation and regeneration to a far lesser extent compared to matrices from neonatal tissues, which are softer. Wow, so this ties in with the talk suggesting that massaging prevents ageing. So get a massage today to relax all that stiffness, and perhaps maintain a softer matrix!

Final talk of the session was delivered by Shivashankar. He suggested that matrix constraints could determine the chromatin packing in the cells. This could be critical to bring together co-regulated genes on different chromosomes into gene clusters. Within these boundary conditions there could be an erasure of the genomic memory of the cells and help transform them on the genetic landscape to cells of another type. This was demonstrated by the growth of fibroblasts on fibronectin presenting rectangular substrates and for a few days lead to the crowding of cells and they began to express E-Cadherin. Finally, they propose to use nuclear features as a biomarker for several diseases in the tissue.

After this session, poster prizes were awarded. Really exciting posters, ranging from broad topics of integrins to cadherins and engineering substrates to genomic data analyses in cancer cells and to unconventional (previously annotated with very different functions!) genes expressed in cancer tissue were recognized.

New Interdisciplinary Connections at the Mechanobiology Meeting-An Undergraduate Perspective

Following a topical link with previous Gordon Conferences targeted at mechanosensitive ion channels, this fall’s Biophysical Society thematic meeting focused on the “Mechanobiology of Proteins and Cells.”  It was held at the Mount Desert Island Biological Laboratory (MDIBL) in Salisbury Cove, ME, and drew a wide range of international investigators interested in a physical understanding of cellular function.  The scientific program maintained an excellent balance between focus and breadth; indeed our research group (consisting of a majority of undergraduate research students) found it to be highly informative, clearly presented, motivational, and inspiring. 

The Burden Research Group at the Mechanobiology Meeting

The Burden Research Group at the Mechanobiology Meeting

Example session topics included biophysical tools for studying the mechanics of proteins and cells, cellular osmoregulation, mechanosensing, ion channel structural biology, cytoplasmic networks, and the cytoskeleton.  Throughout the week, the science was a visual feast, particularly for those with a bend towards imaging technologies that augment and pioneer new discoveries.  An intriguing combination of both in-depth presentations and shorter “poster advertisement” talks were thoughtfully sequenced by the meeting organizers.

Because our youngest team members are studying in the traditional tracks of chemistry, biology, physics, and computer science, this venue provided a particularly poignant opportunity to experience interdisciplinary science at its best.  Even more exhilarating, the general youthfulness of our team made little difference to our interactions with other investigators at the poster sessions, around the dinner table, and at other social functions.  Example testimonials included:

 “Though the most inexperienced of the scientists attending the conference, we were warmly received. The scientists entertained our questions on their research, and many of them had helpful critiques and praises for our research that we presented at the poster sessions.”

 “I was surprised by the ability to connect with any speaker throughout the week.”

 “Poster session discussions with other scientists pursuing similar courses of research to mine gave me the opportunity to brainstorm and collaborate with others to advance our research.”

“I certainly did not expect the degree of camaraderie that I witnessed at the conference. It was so encouraging to see top-notch scientists not only question and imagine together but also affirm each other. The congenial sharing of ideas that I experienced at the conference was both exciting and refreshing.”

Overall, the conference showcased a vibrant and growing field.  With many decades of expansion to come in mechanobiology, our group left with a sense of historical privilege and gratitude that we should have the opportunity to participate at this particular juncture, as the field continues to unfold.

-The Burden Research Group
Wheaton College Chemistry and Biology Departments
Special contributions from Dan Burden, Elese Lau, Olivia Coury, and Hannah Phelps