Intracellular gene transfer in action: Dual transcription and multiple silencings of nuclear and mitochondrial cox2 genes in legumes

The respiratory gene cox2, normally present in the mitochondrion, was previously shown to have been functionally transferred to the nucleus during flowering plant evolution, possibly during the diversification of legumes. To search for novel intermediate stages in the process of intracellular gene transfer and to assess the evolutionary timing and frequency of cox2 transfer, activation, and inactivation, we examined nuclear and mitochondrial (mt) cox2 presence and expression in over 25 legume genera and mt cox2 presence in 392 genera. Transfer and activation of cox2 appear to have occurred during recent legume evolution, more recently than previously inferred. Many intermediate stages of the gene transfer process are represented by cox2 genes in the studied legumes. Nine legumes contain intact copies of both nuclear and mt cox2, although transcripts could not be detected for some of these genes. Both cox2 genes are transcribed in seven legumes that are phylogenetically interspersed with species displaying only nuclear or mt cox2 expression. Inactivation of cox2 in each genome has taken place multiple times and in a variety of ways, including loss of detectable transcripts or transcript editing and partial to complete gene loss. Phylogenetic evidence shows about the same number (3–5) of separate inactivations of nuclear and mt cox2, suggesting that there is no selective advantage for a mt vs. nuclear location of cox2 in plants. The current distribution of cox2 presence and expression between the nucleus and mitochondrion in the studied legumes is probably the result of chance mutations silencing either cox2 gene.

Speech technology in the year 2001.

This paper introduces the session "Technology in the Year 2001" and is the first of four papers dealing with the future of human-machine communication by voice. In looking to the future it is important to recognize both the difficulties of technological forecasting and the frailties of the technology as it exists today--frailties that are manifestations of our limited scientific understanding of human cognition. The technology to realize truly advanced applications does not yet exist and cannot be supported by our presently incomplete science of speech. To achieve this long-term goal, the authors advocate a fundamental research program using a cybernetic approach substantially different from more conventional synthetic approaches. In a cybernetic approach, feedback control systems will allow a machine to adapt to a linguistically rich environment using reinforcement learning.

Speech technology in 2001: new research directions.

Research in speech recognition and synthesis over the past several decades has brought speech technology to a point where it is being used in "real-world" applications. However, despite the progress, the perception remains that the current technology is not flexible enough to allow easy voice communication with machines. The focus of speech research is now on producing systems that are accurate and robust but that do not impose unnecessary constraints on the user. This chapter takes a critical look at the shortcomings of the current speech recognition and synthesis algorithms, discusses the technical challenges facing research, and examines the new directions that research in speech recognition and synthesis must take in order to form the basis of new solutions suitable for supporting a wide range of applications.

The future of voice-processing technology in the world of computers and communications.

This talk, which was the keynote address of the NAS Colloquium on Human-Machine Communication by Voice, discusses the past, present, and future of human-machine communications, especially speech recognition and speech synthesis. Progress in these technologies is reviewed in the context of the general progress in computer and communications technologies.

Detection of neuritic plaques in Alzheimer’s disease by magnetic resonance microscopy

Magnetic resonance microscopy (MRM) theoretically provides the spatial resolution and signal-to-noise ratio needed to resolve neuritic plaques, the neuropathological hallmark of Alzheimer’s disease (AD). Two previously unexplored MR contrast parameters, T2* and diffusion, are tested for plaque-specific contrast to noise. Autopsy specimens from nondemented controls (n = 3) and patients with AD (n = 5) were used. Three-dimensional T2* and diffusion MR images with voxel sizes ranging from 3 × 10−3 mm3 to 5.9 × 10−5 mm3 were acquired. After imaging, specimens were cut and stained with a microwave king silver stain to demonstrate neuritic plaques. From controls, the alveus, fimbria, pyramidal cell layer, hippocampal sulcus, and granule cell layer were detected by either T2* or diffusion contrast. These structures were used as landmarks when correlating MRMs with histological sections. At a voxel resolution of 5.9 × 10−5 mm3, neuritic plaques could be detected by T2*. The neuritic plaques emerged as black, spherical elements on T2* MRMs and could be distinguished from vessels only in cross-section when presented in three dimension. Here we provide MR images of neuritic plaques in vitro. The MRM results reported provide a new direction for applying this technology in vivo. Clearly, the ability to detect and follow the early progression of amyloid-positive brain lesions will greatly aid and simplify the many possibilities to intervene pharmacologically in AD.

Abnormal neurotransmission in mice lacking synaptic vesicle protein 2A (SV2A)

Synaptic vesicle protein 2 (SV2) is a membrane glycoprotein common to all synaptic and endocrine vesicles. Unlike many proteins involved in synaptic exocytosis, SV2 has no homolog in yeast, indicating that it performs a function unique to secretion in higher eukaryotes. Although the structure and protein interactions of SV2 suggest multiple possible functions, its role in synaptic events remains unknown. To explore the function of SV2 in an in vivo context, we generated mice that do not express the primary SV2 isoform, SV2A, by using targeted gene disruption. Animals homozygous for the SV2A gene disruption appear normal at birth. However, they fail to grow, experience severe seizures, and die within 3 weeks, suggesting multiple neural and endocrine deficits. Electrophysiological studies of spontaneous inhibitory neurotransmission in the CA3 region of the hippocampus revealed that loss of SV2A leads to a reduction in action potential-dependent γ-aminobutyric acid (GABA)ergic neurotransmission. In contrast, action potential-independent neurotransmission was normal. Analyses of synapse ultrastructure suggest that altered neurotransmission is not caused by changes in synapse density or morphology. These findings demonstrate that SV2A is an essential protein and implicate it in the control of exocytosis.

Highly efficient germ-line transmission of proviral insertions in zebrafish.

An important technology in model organisms is the ability to make transgenic animals. In the past, transgenic technology in zebrafish has been limited by the relatively low efficiency with which transgenes could be generated using either DNA microinjection or retroviral infection. Previous efforts to generate transgenic zebrafish with retroviral vectors used a pseudotyped virus with a genome based on the Moloney murine leukemia virus and the envelope protein of the vesicular stomatitis virus. This virus was injected into blastula-stage zebrafish, and 16% of the injected embryos transmitted proviral insertions to their offspring, with most founders transmitting a single insertion to approximately 2% of their progeny. In an effort to improve this transgenic frequency, we have generated pseudotyped viral stocks of two new Moloney-based genomes. These viral stocks have titers up to two orders of magnitude higher than that used previously. Injection of these viruses resulted in a dramatic increase in transgenic efficiency; over three different experiments, 83% (110/133) of the injected embryos transmitted proviral insertions to 24% of their offspring. Furthermore, founders made with one of the viruses transmitted an average of 11 different insertions through their germ line. These results represent a 50- to 100-fold improvement in the efficiency of generating transgenic zebrafish, making it now feasible for a single lab to rapidly generate tens to hundreds of thousands of transgenes. Consequently, large-scale insertional mutagenesis strategies, previously limited to invertebrates, may now be possible in a vertebrate.

What does voice-processing technology support today?

This paper describes the state of the art in applications of voice-processing technologies. In the first part, technologies concerning the implementation of speech recognition and synthesis algorithms are described. Hardware technologies such as microprocessors and DSPs (digital signal processors) are discussed. Software development environment, which is a key technology in developing applications software, ranging from DSP software to support software also is described. In the second part, the state of the art of algorithms from the standpoint of applications is discussed. Several issues concerning evaluation of speech recognition/synthesis algorithms are covered, as well as issues concerning the robustness of algorithms in adverse conditions.