R E V I E W A R T I C L EIt is all about resolution Meeting report based upon presentations at the 10th International Global BioMillennium 2006 symposium on molecular cell biology Tbilis
Trang 1R E V I E W A R T I C L E
It is all about resolution
Meeting report based upon presentations at the 10th International Global BioMillennium 2006 symposium on molecular cell biology (Tbilisi, Georgia)
Hermona Soreq1and Alik Honigman2
1 Department of Biological Chemistry, The Hebrew University of Jerusalem, Israel
2 Department of Virology, The Hebrew University of Jerusalem, Israel
Introduction
Improved resolution in time and space is the hallmark
of studies in diverse subfields of the life sciences,
including epigenetics, structural biology, and electron
tomography Combined with dynamic imaging of
immune cells and with data management of cDNA
microarrays, such studies yield reliable synchronization
of cell cycle events, with one goal being the
develop-ment of specific microRNA and protein signatures of
particular tumor cell types, and another being to
fol-low the dynamics of neurite growth in the live brains
of mice
Meeting report
From birth to death of gene products Until recently, epigenetic mechanisms have been equated with the inheritance of chromosomal hetero-chromatin Evidence that transcriptionally active states of chromatin can also be epigenetically main-tained was presented by A Francis Stewart (Dresden) Silencing methylations are epigenetically maintained via methyltransferases that associate with proteins that recognize the methylated epitope and propagate the silent state Together, these observations
Keywords
alternative splicing; cellular imaging;
chromatin; ribosomes; siRNA; ubiquitin
Correspondence
H Soreq, Department of Biological
Chemistry, The Hebrew University of
Jerusalem, Jerusalem 91904, Israel
Fax: +972 2 652 0258
Tel: +972 2 658 5109
E-mail: soreq@cc.huji.ac.il
(Received 23 August 2006, revised 20
November 2006, accepted 11 December
2006)
doi:10.1111/j.1742-4658.2007.05640.x
The 2006 Global BioMillennium Conference took place in Tbilisi, Georgia,
on 13–17 July 2006 The Conference was focused on key aspects of gene expression processes Characteristic of state-of-the-art research in the life sciences, the invited lectures spanned approaches in cell biology, gene expression, and protein function A particular aspect that is special to the BioMillenium series of conferences (this has been the 10th in this series) is the emphasis on new and emerging technologies; the various experts in the subfields that were covered presented what, in their view, should be critical
to enabling future progress
Abbreviations
DAP, death-associated protein; hnRNP, heterogeneous nuclear ribonucleoprotein; miRNA, micro RNA; MMP, matrix metalloprotease.
Trang 2support a polarization model of chromatin that
rein-forces stability
Three post-transcriptional regulation mechanisms
were presented Alternative splicing facilitates large
pro-teomic complexity with a limited number of genes, as
was discussed by Javier F Caceres (Edinburgh, UK)
The trans-acting factors involved include both
serine-arginine (SR) and heterogeneous nuclear
ribonucleo-protein (hnRNP) A⁄ B proteins, with antagonistic
activities influencing different modes of alternative
spli-cing in vivo, affecting tissue-specific or developmental
regulation of gene expression Certain SR and hnRNP
proteins shuttle continuously between the nucleus and
the cytoplasm Interestingly, hnRNP A1 binds to and is
necessary for the processing of a cluster of intronic
microRNAs suggested to act as a human oncogene
Nonsense-mediated mRNA decay selectively degrades
mRNAs harboring premature termination codons In
humans, nonsense-mediated mRNA is linked to
splicing, but in Drosophila and also in Caenorhabditis
elegans, it occurs independently of introns A polarity
effect reduces nonsense-mediated mRNA sensitivity to a
region close to the 3¢-end of the mRNA
As a next step in the study of the regulation of
gene expression, Ada Yonath (Rehovot, Israel)
linked ribosomal architecture with antibiotic action
In all known ribosome structures, an internal
symmet-rical region connects all functional features The
sym-metry relates the RNA backbone to nucleotide
orientation, but shows no sequence homology This
demonstrates the superiority of function over
sequence conservation, suggesting that ribosomes
evolved by gene fusion Antibiotics complexed with
ribosomes from an archaeon that shares properties
with eukaryotes illuminated the structural elements
required for therapeutic effectiveness Structural
bio-logy can hence become a key tool for developing
novel antibiotics
Post-translational regulation was discussed by Aaron
Ciechanover (Haifa, Israel), who argued that the
ubiquitin proteolytic system covers the pathway for
elucidating the basic mechanisms that are pivotal for
drug targeting Degradation of cellular proteins plays
major roles in a multitude of basic pathways during
cell life and death, in both health and disease The
ubiquitin–proteasome pathway involves conjugation of
multiple ubiquitin moieties to the substrate and
subse-quent degradation of the tagged protein, which
involves the downstream 26S proteasome complex and
unknown mechanisms The common thread in all of
these topics is far greater complexity than was
previ-ously perceived, and the need for methods that achieve
the maximum resolution
Imaging genomic and cellular properties Different technologies dealing with real-time imaging
of molecular events, which has become a major tool in biological research, were presented
Hans J Tanke (Leiden, the Netherlands) described cellular imaging of telomere localization and dynamics
in normal cells and in cancer cells using a fluorescently labeled peptide nucleic acid probe with a sequence complementary to telomeric DNA Engineering of fusion proteins of telomere-binding proteins with yel-low fluorescent protein and time-lapse imaging were used to follow the dynamic behavior of telomeres, which were shown to interact dynamically with nuclear bodies
Matthias Gunzer (Braunschweig, Germany) studies cell motility and migration of immune cells, which changes considerably when classical liquid cell culture systems are exchanged for more physiologic environ-ments Mimicking the features of true extracellular tissue can be achieved by embedding the cells in hydra-ted gels of type 1 collagen, or in undisturbed tissues of living animals
Itamar Simon (Jerusalem, Isreal) described a gen-ome-wide analysis of the human cell cycle in primary cells Combining microarray expression data with pre-cise measurements of the culture synchrony at each time point enables deconvolution of the temporal expression signal, yielding coherent, single cell-based expression patterns Microarray and complementary fluorescence-activated cell sorting experiments were used to test a number of arrest methods for normal fibroblast cells Combination of the findings with exist-ing cancer cell cycle data highlights three groups of genes) those that cycle in both cancer and normal cells; those that cycle only in normal cells; and those that cycle only in cancer cells) providing new insights into the transformation process
Continuous exchange of macromolecules between the nucleus and cytoplasm is mediated by nuclear pore complexes, macromolecular assemblies that fuse the inner and outer nuclear membrane and form aqueous channels for translocation Ohad Medalia (Beer-Sheva, Israel) reported on structural analysis of the nuclear pore complex by cryo-electron tomography, exploring transport-active intact nuclei from the slime mold Dictyostelium discoideum Computerized three-dimen-sional classification and averaging provided a refined structure of the nuclear pore complex, enabling the construction of the trajectories of import complexes using gold-labeled substrates
Nuclear export emerges as the major regulatory mechanism of the nuclear accumulation of STAT2, as
Trang 3reported by Hansjo¨rg Hauser (Braunschweig,
Ger-many) In the absence of interferon, STAT2
perma-nently and rapidly shuttles between the cytoplasm and
the nucleus A region in the C-terminus of STAT2
controls its specific export in the absence of interferon
STAT1 also shuttles in the absence of interferon, but
the exchange rate in unstimulated cells is more than 10
times lower In this context Georgyi Los (Madison,
WI, USA) and Marjeta Urh (Madison, WI, USA)
described the use of a fusion between a modified
halo-genase and target proteins to explore protein functions
and dynamics in live cells
Adi Kimchi (Rehovot, Israel) reported on
mole-cular networks involved in programmed cell death
She identified gene products, named DAPs
(death-associated proteins), that differ substantially in their
biochemical properties and intracellular localization
DAP-kinases (DAPk), Ca2+⁄ calmodulin-regulated
and Ser⁄ Thr kinases, activate signaling pathways that
lead to cell death through membrane blebbing and
autophagic cell death Two closely related kinases,
ZIPk and DRP-1, mediate trans-phosphorylation and
subsequent functional activation of ZIPk DAP5 is a
translation initiation factor that directs internal
ribo-some entry site (IRES)-dependent translation under
stress conditions when translation dependent on
5¢ methyl-protected guanosine, designated ‘cap’, is
compromised RNA interference technology serves to
knock down various components of the network
singly or in combination, and validate these
conclu-sions The linking theme in this session highlighted
the need to combine various methodologies in the
search for regulatory events
Approaching tumorigenesis
The challenge of cross-platform analysis of cancer
microarray data was presented by Roland Eils
(Heidel-berg), who uses median rank scores and quantile
dis-cretization to derive numerically comparable measures
of gene expression from different platforms Applied
to six publicly available cancer microarray gene
expres-sion datasets from three pairs of studies, this approach
was used for examining breast cancer, prostate cancer
and acute myeloid leukemia Two leukemia microarray
datasets were further employed to identify important
genes with regard to the biology of leukemia
Import-antly, these could be found in an integrated analysis
but were missed in the single-set analyses
Cross-platform classification of multiple cancer microarray
datasets may yield discriminative gene expression
sig-natures generated by different laboratories and
micro-array technologies
Reuven Agami (Amsterdam, the Netherlands) dem-onstrated the identification of oncogenic microRNAs using a functional genetic screen The number of veri-fied human micro (mi)RNAs is still expanding, but only few have been functionally annotated Agami and colleagues developed a library of vectors expressing the majority of cloned human miRNAs, and created corresponding DNA ‘barcode’ arrays miR-372 and miR-373 were identified, each permitting proliferation and tumorigenesis of primary human cells, neutralizing p53-mediated CDK2 inhibition High-throughput for-mats of miRNA screens will be required to search for other cancer-related functions of miRNAs
The emerging approach of dynamic imaging of pro-tease function in cancer cell invasion was covered by Peter Friedl (Wu¨rzburg, Germany), who combined confocal and multiphoton microscopic imaging of cell dynamics During invasive single-cell migration, the cleavage of individual fiber belts occurs at regions of focal pressure, such as fiber insertions at branching pseudopods or nuclear compression zones These open small degradation tracks, and continuously expanding tubes then become filled with cells and mobile cell masses Pericellular proteolysis is hence a prerequisite for expansive tumor growth, collective invasion and large-scale tissue reshaping In contrast, single-cell dis-semination associated with subtle extracellular matrix remodelling is mechanistically independent of protease function and may be rescued by nonproteolytic escape strategies In this session as well, the complexity of processes and the need for high-resolution strategies were evident
Addressing the brain frontiers Adi Mizrahi (Jerusalem, Israel) reported the use of two-photon microscopy for studying how neuronal dendrites form in adult-born neurons that continuously develop into the olfactory bulb Lentivirus green fluor-escent protein-labeled neurons were used to directly follow dendritic development in vivo Within a single day, neuronal morphology changed dramatically, with both formation and retraction of whole dendritic trees After 10 days, most of the neurons were still migra-ting After 45 days, most granule neurons had comple-ted migration and showed elaborate, complex dendritic trees The dynamics of neuronal development in the intact mammalian brain are hence amenable to further study
Elias Michaelis (Lawrence, KS, USA) covered the establishment of the Glud1 transgenic mouse, which overexpresses the mitochondrial enzyme glutamate dehydrogenase 1 (GLUD1), positioned at the
Trang 4second-to-last enzymatic step in the pathway for biosynthesis
of Glu as a transmitter Transgenic mice suffer losses
in specific neuronal populations, e.g pyramidal
neu-rons of the somatosensory cortex, the CA1 region of
the hippocampus, and motoneurons of the spinal cord,
and show extensive damage to dendrites and axons of
spinal cord motor neurons Importantly, the GLUD1
hippocampus showed higher expression in transgenic
stress response genes, Ca2+-regulating genes, and
genes whose products control cell survival and neurite
growth Neuronal signaling regulated by intracellular
Ca2+ and oxidative stress is hence associated with
raft-like domains in membranes Luciferase-based
reporter bioluminescence assays reported by Keith V
Wood (Madison, WI, USA) offer new methods for
measuring such responses, which may become
import-ant targets for therapeutic intervention against
neuro-degeneration and age-dependent cognitive decline
Anxiety disorders present a major mental health
problem Hermona Soreq (Jerusalem, Israel) discussed
anxiety-induced changes in cholinergic
neurotransmis-sion that modulate motor control over movement,
working memory, and brain-to-body communication
through the neuron–immune system interface,
modify-ing blood cell composition and platelet production
Importantly, the acetylcholinesterase ACHE gene
encodes not one protein, but a combinatorial series of
proteins with indistinguishable enzymatic activity but
with variant N-termini and C-termini, due to
alternat-ive promoter usage and 3¢-alternatalternat-ive splicing These
show distinct nonhydrolytic properties, interact with
variant-specific protein partners, and induce inverse
signaling cascades Specifically, causal involvement of
both butyrylcholinesterase and acetylcholinesterase in
the progression of Alzheimer’s and Parkinson’s
dis-eases anticipates future therapeutic needs for drugs
targeting specific cholinesterases or the corresponding
RNA transcripts
Last, but not least, in this session was Leszek
Kacz-marek (Warsaw, Poland), who discussed extracellular
proteolysis in neuronal plasticity, learning and mem-ory C-Fos and its functional form, the AP-1 transcrip-tion factor, emerged in his studies as the best correlate
of learning processes, especially of the behavioral information Matrix metalloproteinases (MMPs) are pivotal for tissue remodeling in the rodent hippocam-pus, where both matrix metalloproteinase protein-9 (MMP-9) protein and its transcript are associated with
a subset of dendritic spines bearing asymmetric, excita-tory synapses Furthermore, functional inactivation of MMP-9 by means of either gene knockout, or specific chemical inhibitors or TIMP-1, delivered by an adeno-virus vector, affected neuronal plasticity and blocked the late phase of long-term potentiation as well as hip-pocampus-dependent learning Thus, several different proteins were discussed in this session as being relevant for neuronal events characteristic of brain development and aging; however, the gap between cellular and molecular studies appeared to be larger in neuroscience than in tumor-related projects
Acknowledgements
We would like to thank the Promega Corporation of Madison, WI, USA for supporting this meeting
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