Looking back.

In this Perspective, I review my scientific career, which began after I trained in medicine in Montreal and in neurology in Boston. I started in immunology in London with Avrion Mitchison, using antibodies against cell-surface antigens to study the development and functions of mouse T and B cells. The finding that antibody binding causes immunoglobulin on B cells to redistribute rapidly on the cell surface and be endocytosed transformed me from an immunologist into a cell biologist. I moved with Mitchison to University College London, where my colleagues and I used the antibody approach to study cells of the rodent nervous system, focusing on the intrinsic and extrinsic molecular mechanisms that control the development and behavior of myelinating glial cells-Schwann cells and oligodendrocytes. I retired from active research in 2002 and now spend much of my time on scientific advisory boards and thinking about autism.

New routes into the human brain.

The combination of human genetics, animal models, and induced pluripotent stem cells is likely to revolutionize our understanding of neuropsychiatric disorders, leading to new therapies and insights into how the normal human brain works. This is the territory I would explore if I were starting my research career today.

Open questions: what has genetics told us about autism spectrum disorders?

Some of the most interesting questions in biology today, in my view, derive from the real advances in neuropsychiatry that have come largely from human genetics. Research in autism spectrum disorders (ASDs) has been leading the way, mainly because it has become especially well funded and has recently attracted many outstanding scientists. (I must make it clear that I am an outsider in this field, as I have never worked on any neuropsychiatric disorder).

Importance of intrinsic mechanisms in cell fate decisions in the developing rat retina.

Cell diversification in the developing nervous system is thought to involve both cell-intrinsic mechanisms and extracellular signals, but their relative importance in particular cell fate decisions remains uncertain. In the mammalian retina, different cell types develop on a predictable schedule from multipotent retinal neuroepithelial cells (RNECs). A current view is that RNECs pass through a series of competence states, progressively changing their responsiveness to instructive extracellular cues, which also change over time. We show here, however, that embryonic day 16-17 (E16-17) rat RNECs develop similarly in serum-free clonal-density cultures and in serum-containing retinal explants--in the number of times they divide, the cell types they generate, and the order in which they generate these cell types. These surprising results suggest that extracellular signals may be less important than currently believed in determining when RNECs stop dividing and what cell types they generate when they withdraw from the cell cycle, at least from E16-17 onward.

Asymmetric segregation of Numb: a mechanism for neural specification from Drosophila to mammals.

It is a major challenge to understand how the neuroepithelial cells of the developing CNS choose between alternative cell fates to generate cell diversity. In invertebrates such as Drosophila melanogaster and Caenorhabditis elegans, asymmetric segregation of cell-fate determining proteins or mRNAs to the two daughter cells during precursor cell division plays a crucial part in cell diversification. There is increasing evidence that this mechanism also operates in vertebrate neural development and that Numb proteins, which function as cell-fate determinants during Drosophila development, may also function in this way in vertebrates. Recent studies on mouse cortical progenitor cells have provided the strongest evidence yet that this is the case. Here, we review these and other findings that suggest an important role for the asymmetric segregation of Numb proteins in vertebrate neural development.

Generation and characterization of oligodendrocytes from lineage-selectable embryonic stem cells in vitro.

Oligodendrocytes develop from proliferating oligodendrocyte precursor cells (OPCs), which arise in germinal zones, migrate throughout the developing white matter and divide a limited number of times before they terminally differentiate. Thus far, it has been possible to purify OPCs only from the rat optic nerve, but the purified cells cannot be obtained in large enough numbers for conventional biochemical analyses. Moreover, the central nervous system stem cells that give rise to OPCs have not been purified, limiting the ability to study the earliest stages of commitment to the oligodendrocyte lineage. Pluripotent mouse embryonic stem (ES) cells can be propagated indefinitely in culture and induced to differentiate into various cell types. We describe protocols for culture conditions in which neural precursor cells, OPCs, and oligodendrocytes can be efficiently produced from genetically modified ES cells. This strategy should be useful for study of the intracellular and extracellular factors that direct central nervous system stem cells down the oligodendrocyte pathway and influence subsequent oligodendrocyte differentiation. It may also be useful for producing OPCs and oligodendrocytes from human ES cells for cell therapy and drug screening.

Control and maintenance of mammalian cell size: response.

A response to Cooper S: Control and maintenance of mammalian cell size. BMC Cell Biol 2004, 5:35.

A Novel Type of Glial Cell Associated with Nodes of Ranvier in Rat Optic Nerve.

Using Golgi impregnation and intracellular injection of horseradish peroxidase, we show that the adult rat optic nerve contains two distinct types of astrocyte-like glial cells: one has mainly radially oriented processes that terminate on blood vessels or on the pial surface; the other has mainly longitudinally oriented processes that associate with, and often terminate at, nodes of Ranvier, but do not end on blood vessels or the pial surface. The sequence of appearance of the two types of glial cells in the developing nerve, taken together with previous immunocytochemical findings, suggests that these cells may correspond to the two types of astrocytes previously described in cultures of perinatal optic nerve cells-those with mainly radially oriented processes corresponding to type-1 astrocytes and those with mainly longitudinally oriented processes corresponding to type-2 astrocytes. To our knowledge, this is the first description of a class of central nervous system (CNS) glial cell whose processes are primarily associated with nodes of Ranvier.

Adult stem cell plasticity: fact or artifact?

There has been unprecedented recent interest in stem cells, mainly because of the hope they offer for cell therapy. Adult stem cells are an attractive source of cells for therapy, especially in view of the recent claims that they are remarkably plastic in their developmental potential when exposed to new environments. Some of these claims have been either difficult to reproduce or shown to be misinterpretations, leaving the phenomenon of adult stem cell plasticity under a cloud. There are, however, other examples of plasticity where differentiated cells or their precursors can be reprogrammed by extracellular cues to alter their character in ways that could have important implications for cell therapy and other forms of regenerative treatment.

Chromatin remodeling and histone modification in the conversion of oligodendrocyte precursors to neural stem cells.

We showed previously that purified rat oligodendrocyte precursor cells (OPCs) can be induced by extracellular signals to convert to multipotent neural stem-like cells (NSLCs), which can then generate both neurons and glial cells. Because the conversion of precursor cells to stem-like cells is of both intellectual and practical interest, it is important to understand its molecular basis. We show here that the conversion of OPCs to NSLCs depends on the reactivation of the sox2 gene, which in turn depends on the recruitment of the tumor suppressor protein Brca1 and the chromatin-remodeling protein Brahma (Brm) to an enhancer in the sox2 promoter. Moreover, we show that the conversion is associated with the modification of Lys 4 and Lys 9 of histone H3 at the same enhancer. Our findings suggest that the conversion of OPCs to NSLCs depends on progressive chromatin remodeling, mediated in part by Brca1 and Brm.

The orientation of cell division influences cell-fate choice in the developing mammalian retina.

Asymmetric segregation of cell-fate determinants during cell division plays an important part in generating cell diversity in invertebrates. We showed previously that cells in the neonatal rat retina divide at various orientations and that some dividing cells asymmetrically distribute the cell-fate determinant Numb to the two daughter cells. Here, we test the possibility that such asymmetric divisions contribute to retinal cell diversification. We have used long-term videomicroscopy of green-fluorescent-protein (GFP)-labeled retinal explants from neonatal rats to visualize the plane of cell division and follow the differentiation of the daughter cells. We found that cells that divided with a horizontal mitotic spindle, where both daughter cells should inherit Numb, tended to produce daughters that became the same cell type, whereas cells that divided with a vertical mitotic spindle, where only one daughter cell should inherit Numb, tended to produce daughters that became different. Moreover, overexpression of Numb in the dividing cells promoted the development of photoreceptor cells at the expense of interneurons and Müller glial cells. These findings indicate that the plane of cell division influences cell-fate choice in the neonatal rat retina and support the hypothesis that the asymmetric segregation of Numb normally influences some of these choices.

Differences in the way a mammalian cell and yeast cells coordinate cell growth and cell-cycle progression.

BACKGROUND: It is widely believed that cell-size checkpoints help to coordinate cell growth and cell-cycle progression, so that proliferating eukaryotic cells maintain their size. There is strong evidence for such size checkpoints in yeasts, which maintain a constant cell-size distribution as they proliferate, even though large yeast cells grow faster than small yeast cells. Moreover, when yeast cells are shifted to better or worse nutrient conditions, they alter their size threshold within one cell cycle. Populations of mammalian cells can also maintain a constant size distribution as they proliferate, but it is not known whether this depends on cell-size checkpoints. RESULTS: We show that proliferating rat Schwann cells do not require a cell-size checkpoint to maintain a constant cell-size distribution, as, unlike yeasts, large and small Schwann cells grow at the same rate, which depends on the concentration of extracellular growth factors. In addition, when shifted from serum-free to serum-containing medium, Schwann cells take many divisions to increase their size to that appropriate to the new condition, suggesting that they do not have cell-size checkpoints similar to those in yeasts. CONCLUSIONS: Proliferating Schwann cells and yeast cells seem to use different mechanisms to coordinate their growth with cell-cycle progression. Whereas yeast cells use cell-size checkpoints, Schwann cells apparently do not. It seems likely that many mammalian cells resemble Schwann cells in this respect.