A single stem cell can recolonize an embryonic thymus, producing phenotypically distinct T-cell populations

There is much interest in early T-cell development, particularly in relation to the diversification of the T-cell receptor repertoire and the elucidation of the lineage relationships between T-cell populations in the thymus and peripheral lymphoid organs. However, the requirements for the growth of the earliest thymic T-cell precursor in 13-14-day mouse embryo thymus in isolation from the thymic environment are unknown. Proliferation and maturation of such cells are not sustained either in the presence of monolayers of thymic stromal cells or by the addition of interleukin-2 (IL-2), despite the expression of receptors for this growth factor on a proportion of thymocytes displaying the immature Thy 1+ Lyt-2-L3T4- phenotype in the embryonic thymus. In contrast, when maintained within the intact thymic environment in organ cultures, 13-14-day thymic stem cells do show a pattern of surface marker and functional development similar to that seen in vivo, suggesting that short-range growth signals, perhaps necessitating direct contact with organized epithelial cells, are required. We have shown, by exploiting the selective toxicity of deoxyguanosine (dGuo) for early T cells, that this organ culture system can be manipulated to produce alymphoid lobes that can be recolonized from a source of precursors in a transfilter system. We now show that recolonization of alymphoid lobes can also be achieved by association with T-cell precursors in hanging drops, allowing recolonization by exposure to defined numbers of precursors, including a single micromanipulated stem cell. Analysis of T-cell marker expression in these cultures shows that a single thymic stem cell can produce progeny of distinct phenotypes, suggesting that these marker-defined populations are not derived from separate prethymic precursors, but arise within the thymus.     

A single micromanipulated stem cell gives rise to multiple T-cell receptor gene rearrangements in the thymus in vitro

The extensive range of specificities of T-cell receptors is generated, as for immunoglobulins, by rearrangement of genetic information. Much valuable information about rearrangement processes has been inferred by comparing DNA from (monoclonal) lymphoid lines with germ-line DNA and, for B cells, from rearrangements in some Abelson murine leukaemia virus-transformed cell lines. However, because it is difficult to isolate and grow precursor populations, it has not proved possible to study rearrangements occurring in normal untransformed cells in vitro. Here we show that a single T-cell precursor colonizing an alymphoid thymus lobe in organ culture can generate multiple receptor beta-chain gene rearrangements. These observations provide unequivocal evidence for the intra-thymic diversification of the T-cell repertoire. They also offer the possibility of investigating rearrangement and its control in the clonal progeny of a single normal T-cell precursor without the perturbations involved in the use of viral transformation or the production of T-cell hybridomas.     

Mutations in amphiphysin 2 (BIN1) disrupt interaction with dynamin 2 and cause autosomal recessive centronuclear myopathy

Centronuclear myopathies are characterized by muscle weakness and abnormal centralization of nuclei in muscle fibers not secondary to regeneration. The severe neonatal X-linked form (myotubular myopathy) is due to mutations in the phosphoinositide phosphatase myotubularin (MTM1), whereas mutations in dynamin 2 (DNM2) have been found in some autosomal dominant cases. By direct sequencing of functional candidate genes, we identified homozygous mutations in amphiphysin 2 (BIN1) in three families with autosomal recessive inheritance. Two missense mutations affecting the BAR (Bin1/amphiphysin/RVS167) domain disrupt its membrane tubulation properties in transfected cells, and a partial truncation of the C-terminal SH3 domain abrogates the interaction with DNM2 and its recruitment to the membrane tubules. Our results suggest that mutations in BIN1 cause centronuclear myopathy by interfering with remodeling of T tubules and/or endocytic membranes, and that the functional interaction between BIN1 and DNM2 is necessary for normal muscle function and positioning of nuclei.     

Activation in vitro of sequence-specific DNA binding by a human regulatory factor

The human heat-shock factor (HSF) regulates heat-shock genes in response to elevated temperature. When human cells are heated to 43 degrees C, HSF is modified post-translationally from a form that does not bind DNA to a form that binds to a specific sequence (the heat-shock element, HSE) found upstream of heat-shock genes. To investigate the transduction of the heat signal to HSF, and more generally, how mammalian cells respond at the molecular level to environmental stimuli, we have developed a cell-free system that exhibits heat-induced activation of human HSF in vitro. Comparison of HSF activation in vitro and in intact cells suggests that the response of human cells to heat shock involves at least two steps. First, an ATP-independent, heat-induced alteration of HSF allows it to bind the HSE; the temperature at which activation occurs in vitro implies that a human factor directly senses temperature. Second, HSF is phosphorylated. It is possible that similar multi-step activation mechanisms play a role in the response of eukaryotic cells to a variety of environmental stimuli, and that these mechanisms evolved to increase the range and flexibility of the response.     

The Theory of Adaptation of the Nervous System

The explanation and simulation of the natural and artificial intelligence are the central goals of the studies of Neuroscience, Psychology, Artificial Intelligence and Cognitive Science. This paper first gives an introduction to the core topics and approaches in the study. Then, GAF--a general adaptive framework for neural system is proposed. Interdisciplinary discussions around the adaptation of the human nervous system are presented. Rules describing the theory of adaptation of the nervous system are provided.     

Conditionally disordered proteins: bringing the environment back into the fold

For many proteins, biological function requires the folding of the polypeptide chain into a unique and persistent tertiary structure. This review concerns proteins that adopt a specific tertiary structure to function, but are otherwise partially or completely disordered. The biological cue for protein folding is environmental perturbation or minor post-translational modification. Hence, we term these proteins conditionally disordered. Many of these proteins recognize and bind other molecules, and conditional disorder has been hypothesized to allow for more nuanced control and regulation of binding processes. However, this remains largely unproven. The sequences of conditionally disordered proteins suggest their propensity to fold; yet, under the standard laboratory conditions, they do not do so, which may appear surprising. We argue that the surprise results from the failure to consider the role of the environment in protein structure formation and that conditional disorder arises as a natural consequence of the marginal stability of the folded state.     

Importance of IL-2 receptors in intra-thymic generation of cells expressing T-cell receptors

During development, lymphoid stem cells migrate into the thymic rudiment where they proliferate, rearrange their antigen receptor genes and become differentiated into functionally mature T cells. At present, the regulation of these processes is poorly understood, although recent studies have shown that early fetal and adult immature thymocytes express receptors for the T-cell growth factor, interleukin-2 (IL-2). We now present direct evidence that IL-2 receptors have a function in intra-thymic development by demonstrating that proliferation and the generation of cells expressing the T-cell antigen receptor (alpha beta TCR), which is responsible for the recognition of antigens in the context of MHC, are inhibited when antibodies to IL-2 receptors are added to fetal thymus organ cultures. The inhibition is specific in that it does not affect pre-thymic stem cells and can be partially reversed by addition of exogenous recombinant IL-2.