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High-grade serous carcinomas (HGSC) make up the majority of ovarian cancer cases. Unfortunately, they have the lowest survival rates. HGSC is a tumor type that occurs primarily in the ovaries and spreads throughout the abdominal cavity. Most patients are diagnosed with late-stage disease that has already spread. Until recently, therapy has been limited to surgery and traditional chemotherapeutic agents. A systematic examination of the tumor and surrounding tissue — particularly normal cells called fibroblasts — has revealed a new treatment target that could potentially prevent the rapid dissemination and poor prognosis associated with high-grade serous carcinoma.

In close collaboration with Fabian Coscia and Matthias Mann, from the Max Planck Institute of Biochemistry in Munich and University of Copenhagen, researchers profiled the expression of more than 5,000 proteins in both normal and cancerous tissues derived from minute amounts of patient biobank material. “When we then got our first data, we were fascinated to find that the metastatic stroma was characterized by a highly conserved protein signature, as opposed to the cancer cells”, adds Fabian Coscia, postdoctoral researcher in Matthias Mann’s group and one of the two first-authors of the study. As these metastatic changes were seen in all of their analyzed patients, the team then went on to understand its functional role during metastasis with the ultimate goal to find novel therapeutic targets.

Indeed, they discovered a metabolic enzyme, nicotinamide N-methyltransferase (NNMT), highly expressed in the stroma surrounding metastatic cancer cells. The researchers found that NNMT causes widespread gene expression changes in the tumor stroma, converting normal fibroblasts to cancer-associated fibroblasts that support and accelerate tumor growth. Stromal NNMT expression encouraged ovarian cancer migration, proliferation, growth and metastasis. It was associated with poor clinical outcomes in patients.

They also found that inhibition of NNMT activity may be able to reduce or even reverse many of the tumor-promoting effects of cancer-associated fibroblasts. This suggests, they note, that the stroma should be explored as a new treatment target. Coscia, co-first author on the manuscript who led the proteomics analysis, added that “this method may be used to discover other proteins that are important for metastasis and to identify early changes during disease development.”

“When we put it all together,” Lengyel added, “it gave us exciting results. We have linked high-end technology including proteomics and metabolomics to functional analysis to improve our understanding of the stroma."

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Hopfler, M., Kern, M.J., Straub, T., Prytuliak, R., Habermann, B.H., Pfander, B., and Jentsch, S.
EMBO J, 2019, [Epub ahead of print].
(IMPRS-LS students are in bold)
doi: 10.15252/embj.2018100368

Slx5/Slx8-dependent ubiquitin hotspots on chromatin contribute to stress tolerance

Chromatin is a highly regulated environment, and protein association with chromatin is often controlled by post-translational modifications and the corresponding enzymatic machinery. Specifically, SUMO-targeted ubiquitin ligases (STUbLs) have emerged as key players in nuclear quality control, genome maintenance, and transcription. However, how STUbLs select specific substrates among myriads of SUMOylated proteins on chromatin remains unclear. Here, we reveal a remarkable co-localization of the budding yeast STUbL Slx5/Slx8 and ubiquitin at seven genomic loci that we term "ubiquitin hotspots". Ubiquitylation at these sites depends on Slx5/Slx8 and protein turnover on the Cdc48 segregase. We identify the transcription factor-like Ymr111c/Euc1 to associate with these sites and to be a critical determinant of ubiquitylation. Euc1 specifically targets Slx5/Slx8 to ubiquitin hotspots via bipartite binding of Slx5 that involves the Slx5 SUMO-interacting motifs and an additional, novel substrate recognition domain. Interestingly, the Euc1-ubiquitin hotspot pathway acts redundantly with chromatin modifiers of the H2A.Z and Rpd3L pathways in specific stress responses. Thus, our data suggest that STUbL-dependent ubiquitin hotspots shape chromatin during stress adaptation.

A collaboration between Georgia Tech and the Max Planck Institute of Neurobiology (MPIN) has received a grant of $750,000 over three years from the Human Frontier Science Program (HFSP). The award will allow research on the molecular and genetic encoding of complex behaviors.

The team is led by Georgia Tech’s J. Todd Streelman and MPIN’s Herwig Baier. Streelman is a professor in, and the chair of, the Georgia Tech School of Biological Sciences. Baier is professor and director at the Max Planck Institute of Neurobiology. “It remains incredibly difficult to identify the cellular basis and the genetic variants underlying complex behavior,” Streelman says. “Understanding how behavior is encoded requires solving a dual problem involving neurodevelopment and circuit function.”

To find answers, Streelman and Baier will develop a model system to chart the complex path from genome to brain to behavior in cichlid fish from Lake Malawi. Male cichlid fish build bowers to attract females for mating. The bowers are either pits, which are depressions in the sand, or castles, which look like volcanoes. Each type corresponds to a specific behavior encoded in a fish strain.

When the two strains mate, their male offspring display a remarkable behavior: First they construct a pit then a castle. This behavior indicates that a single brain containing two genomes can produce each behavior in succession. Moreover, gene expression in the brain is biased toward the pit variant of the genome – or pit allele -- when the fish are digging pits and toward the castle allele when they are building castles. “This phenomenon offers the chance to identify both the genome regulatory logic and the neural circuitry underlying complex behavior in one sweep,” Baier says.

Streelman’s group will use single-cell RNA sequencing to pinpoint the cell populations that mediate context-dependent, allele-specific expression in male bower builders. Baier’s team will use genome editing and optogenetic tools to manipulate particular neurons in the brains of behaving bower builders.

The award is one of only 25 made from a total of 654 letters of intent HFSP received from around the world. HFSP provides funding for frontier research in the life sciences. The highly competitive program is implemented by the International Human Frontier Science Program Organization, based in Strasbourg, France.

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Lemke, S.B., Weidemann, T., Cost, A.L., Grashoff, C., and Schnorrer, F.
PLoS Biol, 2019, 17, e3000057.
doi: 10.1371/journal.pbio.3000057

A small proportion of Talin molecules transmit forces at developing muscle attachments in vivo.

Cells in developing organisms are subjected to particular mechanical forces that shape tissues and instruct cell fate decisions. How these forces are sensed and transmitted at the molecular level is therefore an important question, one that has mainly been investigated in cultured cells in vitro. Here, we elucidate how mechanical forces are transmitted in an intact organism. We studied Drosophila muscle attachment sites, which experience high mechanical forces during development and require integrin-mediated adhesion for stable attachment to tendons. Therefore, we quantified molecular forces across the essential integrin-binding protein Talin, which links integrin to the actin cytoskeleton. Generating flies expressing 3 Förster resonance energy transfer (FRET)-based Talin tension sensors reporting different force levels between 1 and 11 piconewton (pN) enabled us to quantify physiologically relevant molecular forces. By measuring primary Drosophila muscle cells, we demonstrate that Drosophila Talin experiences mechanical forces in cell culture that are similar to those previously reported for Talin in mammalian cell lines. However, in vivo force measurements at developing flight muscle attachment sites revealed that average forces across Talin are comparatively low and decrease even further while attachments mature and tissue-level tension remains high. Concomitantly, the Talin concentration at attachment sites increases 5-fold as quantified by fluorescence correlation spectroscopy (FCS), suggesting that only a small proportion of Talin molecules are mechanically engaged at any given time. Reducing Talin levels at late stages of muscle development results in muscle-tendon rupture in the adult fly, likely as a result of active muscle contractions. We therefore propose that a large pool of adhesion molecules is required to share high tissue forces. As a result, less than 15% of the molecules experience detectable forces at developing muscle attachment sites at the same time. Our findings define an important new concept of how cells can adapt to changes in tissue mechanics to prevent mechanical failure in vivo.