Sadaf Shadan

The ubiquitin system

461 Principles of ubiquitin and SUMO modifications in DNA repair S. Bergink & S. Jentsch The destruction of proteins is as important as their synthesis for the maintenance of protein homeostasis in cells. In eukaryotes, the ubiquitin–proteasome system is responsible for most of this protein degradation: the small protein ubiquitin acts as a death warrant, tagging and targeting other proteins to the large proteolytic chamber of the proteasome. The discovery in the 1970s that certain proteins are ubiquitylated before degradation was awarded the 2004 Nobel Prize in Chemistry. It is now known that ubiquitin-mediated destruction plays a crucial part in cell-cycle regulation, DNA repair, cell growth and immune function, as well as in hormone-mediated signalling in plants. More recently, ubiquitin has been shown to have numerous non-proteolytic functions, including involvement in vesicular trafficking pathways, regulation of histone modification and viral budding. Given the central role of the ubiquitin system in diverse cellular processes, it is not surprising that its dysfunction contributes to cancer and to neurodegenerative and immunological disorders. An understanding of the ubiquitin system is therefore important in devising treatments for such diseases. With topics as diverse as the origin of the ubiquitin system and cancer therapy targeting the ubiquitin pathway, this Insight provides both an introduction and an update to the most topical themes in ubiquitin research. We are pleased to acknowledge the financial support of Hybrigenics, ITI Life Sciences and Scil Proteins in producing this Insight. As always, Nature retains sole responsibility for editorial content and peer review.

Frontiers in biology

The Nature Insight ‘Frontiers in biology’ aims to cover timely and important developments in biology, ranging from the subcellular to the organismal level, and including molecular mechanisms and biomedicine. The collection begins with a Review on metastasis, which remains a significant challenge in the treatment of cancer. Metastases in some organs are less responsive to therapy than those in others. Joan Massagué and Anna Obenauf discuss the processes that underlie the colonization of various organs by cancer cells that escape from the primary tumour and enter the circulatory system. Improved understanding of these processes could facilitate the early diagnosis of metastases and lead to new therapeutic interventions. Next, Michael Karin and Hans Clevers explore the cellular and molecular mechanisms of protective inflammation in tissue repair and regeneration, with a focus on the intestines and the liver. The positive effects of inflammation play an essential part in the restoration of tissue homeostasis, and offer protection from chronic inflammation, fibrosis and cancer. Endothelial cells form the lining of blood vessels, but they also function in the development, growth and regeneration of organs through their release of paracrine signals known as angiocrine factors. Shahin Rafii and colleagues examine the emerging role of endothelial cells as signalling centres that orchestrate organ regeneration, morphogenesis and homeostasis. Cells counteract endoplasmic reticulum stress — the abnormal accumulation of misfolded and unfolded proteins of the secretory pathway — by activating a homeostatic process called the unfolded protein response. Miao Wang and Randal Kaufman summarize our current understanding of this field at the molecular and cellular levels, as well as in relation to disease. Resistance to antibiotics poses one of the most pressing threats to modern health care. Eric Brown and Gerard Wright consider the history and use of antibiotics, the factors that have led to the alarming rise in resistant pathogens, and the prospects for overcoming hurdles that hold back antibiotic discovery in the twenty-first century.

Obesity: Fat chance.

nasty, self-sharpening scissor blades. The largest of these fishes equalled the size of a great white shark and must have been formidable predators, but most were less than a metre in length. Working with three-dimensional placoderm fossils from Gogo in Western Australia, Long and colleagues have discovered specimens of three different placoderms, Materpiscis, Austroptyctodus and Incisoscutum, that contain minute but perfectly preserved armour plates of the same species in their body cavities. When first discovered, they were thought to be stomach contents. But the plates show no bite marks or etching by stomach acids, and are not mixed with bones from other species; they are the remains of unborn embryos. In Materpiscis, a curving tubular structure associated with one of them has been interpreted as an umbilical cord. Embryos in the body cavity imply internal fertilization. It was noted long ago that ptycto donts, the placoderm subgroup to which Materpiscis and Austroptyctodus belong, have sexually dimorphic pelvic fins, somewhat like the ‘claspers’ used for internal fertilization in sharks. Arthrodires, the placoderm group that includes Incisoscutum, lack the sexually dimorphic external bones present on the pelvic fins of ptyctodonts. However, Long et al. argue that the partially preserved internal fin skeletons of their specimens indicate a shark-like structure, probably implying sexual dimorphism and internal fertilization. Ptyctodonts and arthrodires seem to be closely related, and so internal fertilization, and possibly live birth of young, are probably shared features retained from their common ancestor. The living jawed vertebrates, or gnathostomes, fall into two groups, the Chondrichthyes and the Osteichthyes (Fig. 1). The Chondrichthyes (sharks, rays and ratfishes) all have internal fertilization, and many give birth to live young, whereas the ancestral condition for the Osteichthyes (ray-finned fishes, lobe-finned fishes and land vertebrates) is to spawn small eggs that are fertilized externally. Live-bearers tend to produce much fewer young than external spawners and have lower potential rates of population growth. This contrast in reproduction puts a new perspective on the ecology of the Gogo environment, a tropical reef , where a wide diversity of placoderms coexisted with lungfishes and primitive ray-finned fishes that were probably externally fertilizing spawners. It is also interesting to note that the extinction of the placoderms at the end of the Devonian was followed by a major diversification of chondrichthyans. But it is to the study of gnathostome interrelationships that the discoveries of Long et al. may prove to be most pertinent. Ideas about the origin of gnathostomes are currently in a state of flux. For much of the twentieth century, placoderms were regarded as relatives or possibly ancestors of chondrichthyans, partly because they seemed to use internal fertilization. But recently the majority view has placed them in the gnathostome stem group — that is, the common ancestral lineage of the living jawed vertebrates. A new analysis by Brazeau suggests that placoderms may not be a natural group at all, but a ‘paraphyletic array’ spread out along the gnathostome stem (Fig. 1a; contrast with Fig. 1b). If that is correct, placoderms become extremely informative about the origin of jawed vertebrate morphology. This is where the evidence for internal fertilization and livebearing in placoderms becomes important. The ancestral mode of reproduction for osteichthyans seems to be external fertilization. The distribution of live-bearing among living vertebrates strongly suggests that internally fertilizing live-bearers are unlikely to give rise to externally fertilizing spawners, so we would not expect the osteichthyan stem lineage, or the gnathostome stem lineage below it, to contain a segment characterized by live-bearing. Brazeau’s analysis places the ptyctodonts and arthrodires as successive branches off the gnathostome stem, implying the existence of such a segment unless the two groups have evolved live-bearing independently. However, only a minor change in the tree would be needed to join ptyctodonts and arthrodires together in a clade (that is, forming a single side branch), and thus make the offending stem segment disappear. A more important question is whether the most primitive placoderms, such as the antiarchs (bottom-feeding fishes with armoured pectoral fins), were also live-bearers, because this would undermine the case for the placoderms forming a paraphyletic segment of the gnathostome stem. Long and colleagues argue that the antiarchs had external fertilization. They lack pelvic fins altogether, and fossils have been found of freeliving juveniles that are small and undeveloped enough to correspond to the embryos of Materpiscis, Austroptyctodus and Incisoscutum. It may thus be that both internal fertilization and live-bearing evolved within the placoderms. Perhaps this was a unique innovation in one placoderm clade. Alternatively, could some placoderms be stem gnathostomes and others, those with internal fertilization, stem chondrichthyans? Possibly, but this conflicts with new evidence that the acanthodians (vaguely shark-like fishes, with fin spines and tiny scales, which became extinct about 250 million years ago) form a paraphyletic array encompassing the bases of the chondrichthyan and osteichthyan lineages (Fig. 1a). The tangled skein of jawed-vertebrate origins continues to challenge researchers. But discoveries such as the placoderm embryos of Gogo are giving us the tools to gradually untangle it — as well as showing us intimate glimpses of life in a lost world. ■

Developmental genetics: Time for teeth

could be important for the formation and evolution of fracture lances. The study of how materials fail is crucial to our ability to engineer better ones — for example, to create lighter, stronger and tougher materials. The emergence of instability fracture mechanisms such as those studied by Pons and Karma tend to make it more difficult for cracks to spread because they increase the energy required for a crack to move forwards, thereby enhancing a material’s overall resistance to catastrophic failure. The design of novel materials by deliberately invoking instability fracture mechanisms — perhaps through the creation of material structures that induce local combined tension–tear loading — could provide a powerful strategy to increase a structure’s resistance to failure without the need to introduce additional material components, instead relying solely on structural changes. An intriguing direction in which Pons and Karma’s idea could be taken is the study of failure in biological materials — for example, spider silk, nacre and bone. In these materials, better resistance to fracture is achieved not through the addition of stronger materials but rather by reliance on a hierarchy of structures, in which each hierarchical level has its own fracture mechanism. The synergistic effect of fracture mechanisms at all levels, attained through a seamless merger of structure and material, achieves an overall performance that vastly exceeds that of each individual level. Although we rely on strong element bonding in the design of most engineered materials — such as covalent bonds in glass or polymers, or metallic bonds in steel — bonding in biological materials is often weaker. For example, spider silk is one of the strongest materials known (stronger than steel), yet its strength lies in the cooperation of extremely weak hydrogen bonding between protein molecules. The effective use of multiple structural levels enables biological materials such as silks to achieve high performance without relying on strong bonding. From a slightly different viewpoint, the breakdown of biological materials’ capacity to effectively withstand fracture can lead to injury and disease, as observed, for example, in osteogenesis imperfecta (brittle-bone disease). As such, the study of fracture could also help us to understand the mechanisms that underlie severe diseases and perhaps provide new pathways for treatment. Pons and Karma’s work shows that a computational approach, validated by experiment, is a powerful tool in explaining fundamental issues in materials failure. The emergence of computational materials models that involve multiple scales, from the atomic to the macroscopic, holds great promise in elucidating the complex facets of how materials fail, and could lead to exciting breakthroughs in our understanding of material breakdown in engineering, geology and biology. ■

Plant biology: scent of a rose.

resistance to cold and disease, they make up for with repeated blooming and a distinctive fragrance. It isn’t surprising, then, that plant breeders, fascinated with this ancient rose family, brought it to Europe in the nineteenth century to generate the now popular hybrid tea roses, among many others. But how did the distinctive scent of Chinese roses evolve? Gabriel Scalliet and colleagues now provide the answer (G. Scalliet et al. Proc. Natl Acad. Sci. USA 105, 5927–5932; 2008). Unlike their European counterparts, the main scent component of Chinese roses and their descendants is dimethoxytoluene. In the final steps of its formation, two methyl groups are sequentially added to a precursor molecule in reactions catalysed by enzymes called OOMT1 and OOMT2. These enzymes are almost identical in amino-acid sequence, yet they target different substrates. Scalliet and colleagues pinpoint the crucial amino-acid residues of the enzymes that confer substrate specificity: a tyrosine in OOMT1 and a phenylalanine in OOMT2. Swapping these single residues between the two enzymes switched their substrate specificity, most probably by changing the steric and hydrophobic properties of their substrate-binding sites, where these residues reside. Moreover, OOMT2 could methylate the target of OOMT1 in vitro, albeit with lower efficiency. Two nearly identical enzymes with nearly identical activities, that can both perform similar reactions, although together they are more efficient than alone — these are hallmarks of the products of duplicated genes, which arise when a gene doubles in number and one of the copies then undergoes a minor mutation. Indeed, Scalliet et al. show that, although all of the 13 European rose species they examined carry an OOMTlike gene, only the Chinese species, which are evolutionarily younger, carry two types of this gene. The authors’ further phylogenetic analysis strongly hints that, in Chinese species, the second OOMT gene arose through a duplication event. Roses are not unique in acquiring new functions through gene duplication; this is thought to be a fundamental mechanism for generating diversity between and within organisms. But it isn’t always possible to trace such changes, which makes Scalliet and colleagues’ work of interest not only to plant biologists but also to evolutionary scientists. Sadaf Shadan ERα-associated demethylase enzyme LSD1, which does not influence interchromosomal gene interaction — is required for the subsequent colocalization of the gene pairs with nuclear speckles (structures composed of high concentrations of specific proteins involved in messenger-RNA splicing), thereby probably leading to mRNA maturation (Fig. 1). Transcriptional coactivators are multiprotein complexes that, collectively, have diverse enzymatic activities. So it is tempting to speculate that, together with LSD1, distinct enzymes of a coactivator complex mediate actin polymerization (which might facilitate gene movement within the nucleus), interchromatin interactions and chromatin interface with transcription ‘factories’ — structures that generate mRNAs at higher rates. Ultimately, it is likely that coactivators and other receptor-associated proteins work together to orchestrate the formation of an ERα-activated transcriptional supercomplex comprising Figure 1 | Rambling genes. a, Nunez et al. find that, in the absence of oestrogen, two genes (TFF1 and GREB1) that are activated by this hormone and found on different chromosomes, reside in different locations within the nucleus. b, On exposure to oestrogen, these genes make interchromatin contacts, facilitated by the nuclear receptor ERα, transcriptional coactivators (CoAs), actin and molecular motor proteins. c, Subsequently, the gene pair interacts with a nuclear speckle in an LSD1-dependent manner, creating a multiprotein complex that might act as a transcriptional hub for DNA transcription into mRNA. a b c

Lipids in health and disease

Background: Higher concentrations of serum lipids and apolipoprotein B100 (apoB) are major individual risk factors of atherosclerosis and coronary heart disease. Therefore ameliorative effects of food components against the diseases are being paid attention in the affluent countries. The present study was undertaken to investigate the effect of taurine on apoB secretion and lipid metabolism in human liver model HepG2 cells. Results: The results demonstrated that an addition of taurine to the culture media reduces triacylglycerol (TG)-mass in the cells and the medium. Similarly, cellular cholesterol-mass was decreased. Taurine inhibited the incorporation of [14C] oleate into cellular and medium TG, suggesting the inhibition of TG synthesis. In addition, taurine reduced the synthesis of cellular cholesterol ester and its secretion, suggesting the inhibition of acyl-coenzyme A:cholesterol acyltransferase activity. Furthermore, taurine reduced the secretion of apoB, which is a major protein component of very low-density lipoprotein. Conclusion: This is a first report to demonstrate that taurine inhibits the secretion of apoB from HepG2 cells.

Behavioural neuroscience: Hare-brained flies

It wasn’t always like this. Protection strategies simply weren’t an option in the early days of organic chemistry, because many of the protecting groups widely used today were developed only in the past 50 years. Accordingly, some exemplary syntheses were devised that do not use protecting groups. For example, a useful method was developed in the early 1900s for converting sugars known as pentoses into larger sugars called hexoses. Pentoses contain many reactive hydroxyl (OH) groups that chemists today would almost certainly protect. This three-step method has been invaluable for making sugars that are difficult to obtain from natural sources. More complex syntheses have also been reported. In 1957, a nine-step synthesis of muscarine — a natural product that mimics the action of certain neurotransmitters — did not use a single protecting group. Another approach for avoiding protection strategies is to imitate biochemical routes found in nature. A landmark synthesis of this type was reported by Robert Robinson in 1917; Robinson went on to win the Nobel Prize in Chemistry partly in recognition of this work. He prepared tropinone — a synthetic precursor of the drug atropine — in one step from simple starting materials, without protecting groups. This is considered to be an early example of a biomimetic cascade sequence, in which an initial reaction triggers a defined chain of other reactions, like dominoes toppling in a line. An extreme version of the biomimetic approach uses enzymes to mediate organic reactions. Enzymes have evolved to work with naturally available molecules that clearly do not incorporate synthetic protecting groups. But the early examples of syntheses free of protecting groups are not the norm. As time passed, the number of organic synthetic methods and chemical reagents expanded markedly. This led to an increased use of protecting groups in multi-step reaction sequences — so much so that this approach is hardly questioned today, and potentially interesting reactions that depend on the innate reactivity of unprotected molecules might be missed. So Baran and co-workers have gone back to basics. They describe synthetic routes to structurally complex compounds: ambiguine H and other related molecules that have been isolated from marine organisms. Each molecule is beautifully constructed by the authors without the use of any protecting groups. A good example of their approach is a step in which an aromatic, nitrogen-containing compound (an indole) reacts with another molecule known as a terpene (Fig. 1), mediated by a strong base and a copper salt. This intriguing reaction relies on the presence of a hydrogen atom attached to the nitrogen of the indole fragment. But this hydrogen would not have been present if the nitrogen had been capped with a protecting group. After the introduction of an isonitrile group (R–N + ≡C into the terpene structure, the authors went on to append a hydrocarbon fragment (a prenyl group; Fig. 1) using an unorthodox reaction that relies again on the unprotected indole nitrogen, and on the innate reactivity of the isonitrile functional group. Remarkably, the syntheses of ambiguine H and related molecules required only 7–10 steps, in contrast to the 20–25 steps for previous routes to these syntheses that used protecting groups. This study by Baran and co-workers shows that complex natural products can be synthesized more efficiently by reducing the use of protecting groups. But there are far greater implications of this work — the authors’ strict avoidance of such groups will inspire new chemistry by exposing the intrinsic reactivity of chemical reagents and reactive intermediates. Further work on chemical synthesis using ‘undressed’ molecules should lead to many more advances and innovative developments in organic synthesis. And it might let chemists rest easy at night, safe in the knowledge that years of lab work won’t be destroyed by recalcitrant protecting groups. ■