RESUMEN
BACKGROUND: Recent studies highlight the role of endoplasmic reticulum (ER) stress and aberrant protein degradation in the pathogenesis of neurodegenerative disorders. Herp which is encoded by the HERPUD 1 (homocysteine-inducible, endoplasmic reticulum stress-inducible, ubiquitin-like domain member 1) gene is a stress-response protein localized in the ER membrane of neurons and other cell types. Herp has been suggested to improve ER-folding, decrease ER protein load, and participate in ER-associated degradation (ERAD) of proteins. METHODS: Based on microarray expression profiling results we have predicted an increased expression of HERPUD1 in the substantia nigra of Parkinson's disease (PD) patients. We have now used brain tissue of some of the same and additional cases of sporadic PD to localize Herp mRNA and protein in individual cell types. RESULTS: We found expression of Herp in neurons and in glial cells including astrocytes. These findings were corroborated by in situ hybridization. Accumulation of Herp protein was also detected in the core of Lewy bodies suggesting a role in their formation. Hierarchical clustering analysis identified TWINKLE (PEO1) as the gene whose expression profile was most similar to that of Herp across the PD cohort. CONCLUSIONS: The nigral glial cells that expressed Herp at a high level resembled TUNEL-positive glia. While some of these cells likely undergo degeneration, the strong up-regulation of Herp in glia could help to explain the inflammation-like changes observed in PD ("neuroinflammation") as it has been shown that the unfolded protein response serves as an important regulator of inflammatory genes in other organs.
Asunto(s)
Cuerpos de Lewy/metabolismo , Proteínas de la Membrana/metabolismo , Enfermedad de Parkinson/metabolismo , Sustancia Negra/metabolismo , Anciano , Anciano de 80 o más Años , Astrocitos/metabolismo , Encéfalo/metabolismo , Estudios de Cohortes , ADN Helicasas/genética , ADN Helicasas/metabolismo , Femenino , Expresión Génica , Humanos , Masculino , Proteínas de la Membrana/genética , Persona de Mediana Edad , Proteínas Mitocondriales , Modelos Neurológicos , Neuroglía/metabolismo , Neuronas/metabolismo , Enfermedad de Parkinson/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo , Regulación hacia ArribaRESUMEN
DnaJ/Hsp40 chaperones determine the activity of Hsp70s by stabilizing their interaction with substrate proteins. We have predicted, based on the in silico analysis of a brain-derived whole-genome transcriptome data set, an increased expression of DnaJ/Hsp40 homologue, subfamily B, member 6 (DnaJB6) in Parkinson's disease (PD; Moran et al. [2006] Neurogenetics 7:1-11). We now show that DnaJB6 is a novel component of Lewy bodies (LBs) in both PD substantia nigra and PD cortex and that it is strongly up-regulated in parkinsonian astrocytes. The presence of DnaJB6 in the center of LBs suggests an early and direct involvement of this chaperone in the neuronal disease process associated with PD. The strong concomitant expression of DnaJB6 in astrocytes emphasizes the involvement of glial cells in PD and could indicate a route for therapeutic intervention. Extracellular alpha-synuclein originating from intravesicular alpha-synuclein is prone to aggregation and the potential source of extracellular aggregates (Lee [2008] J. Mol. Neurosci. 34:17-22). The observed strong expression of DnaJB6 by astrocytes could reflect a protective reaction, so reducing the neuronal release of toxic alpha-synuclein and supporting the astrocyte response in PD might limit the progression of the disease process.
Asunto(s)
Astrocitos/metabolismo , Astrocitos/patología , Proteínas del Choque Térmico HSP40/metabolismo , Cuerpos de Lewy/metabolismo , Chaperonas Moleculares/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Trastornos Parkinsonianos/patología , Regulación hacia Arriba/fisiología , Anciano , Anciano de 80 o más Años , Femenino , Proteínas del Choque Térmico HSP40/genética , Humanos , Masculino , Chaperonas Moleculares/genética , Proteínas del Tejido Nervioso/genética , Cambios Post Mortem , ARN Mensajero/metabolismo , alfa-Sinucleína/metabolismoRESUMEN
The pathological basis of dementia and visual hallucinations in Parkinson's disease (PD) is not yet fully understood. To investigate this further we have conducted a clinico-pathological study based on 30 post-mortem PD brains. PD cases were stratified into groups according to clinical characteristics as follows: (1) cognitively intact (n=9); (2) cases with severe dementia and visual hallucinations (n=12); (3) cases with severe dementia and no visual hallucinations (n=4); and (4) cases with severe visual hallucinations and no dementia (n=5). The extent of alpha-synuclein (alphaSyn), tau and amyloid beta peptide (Abeta) deposition was then examined in the CA2 sector of the hippocampus and in neocortical and subcortical areas known to subserve cognitive function. We find that dementia in PD is significantly associated with alphaSyn in the anterior cingulate gyrus, superior frontal gyrus, temporal cortex, entorhinal cortex, amygdaloid complex and CA2 sector of the hippocampus. Abeta in the anterior cingulate gyrus, entorhinal cortex, amygdaloid complex and nucleus basalis of Meynert is also associated with dementia as is tau in the CA2 sector of the hippocampus. alphaSyn burden in the amygdala is strongly related to the presence of visual hallucinations but only in those PD cases with concomitant dementia. Statistical analysis revealed that alphaSyn burden in the anterior cingulate gyrus could differentiate demented from non-demented PD cases with high sensitivity and specificity. We conclude that alphaSyn in limbic regions is related to dementia in PD as well as to visual hallucinations when there is an underlying dementia.
Asunto(s)
Demencia/patología , Alucinaciones/patología , Sistema Límbico/patología , Enfermedad de Parkinson/patología , Adulto , Anciano , Anciano de 80 o más Años , Péptidos beta-Amiloides/metabolismo , Análisis de Varianza , Demencia/etiología , Femenino , Alucinaciones/etiología , Alucinaciones/metabolismo , Humanos , Sistema Límbico/metabolismo , Masculino , Persona de Mediana Edad , Enfermedad de Parkinson/complicaciones , Enfermedad de Parkinson/metabolismo , Cambios Post Mortem , alfa-Sinucleína/metabolismo , Proteínas tau/metabolismoRESUMEN
The identification of mutations that cause familial Parkinson's disease (PD) provides a framework for studies into pathways that may be perturbed also in the far more common, non-familial form of the disorder. Following this hypothesis, we have examined the gene regulatory network that links alpha-synuclein and parkin pathways with dopamine metabolism in neuropathologically verified cases of sporadic PD. By means of an in silico approach using a database of eukaryotic molecular interactions and a whole genome transcriptome dataset validated by qRT-PCR and histological methods, we found parkin and functionally associated genes to be up-regulated in the lateral substantia nigra (SN). In contrast, alpha-synuclein and ubiquitin carboxyl-terminal hydrolase L1 (UCHL1) gene expression levels were significantly reduced in both the lateral and medial SN in PD. Gene expression for Septin 4, a member of the GTP-binding protein family involved in alpha-synuclein metabolism was elevated in the lateral parkinsonian SN. Additionally, catalase and mitogen-activated protein kinase 8 and poly(ADP-ribose) polymerase family member 1 (PARP1) known to function in DNA repair and cell death induction, all members of the dopamine synthesis pathway, were up-regulated in the lateral SN. In contrast, two additional PD-linked genes, glucocerebrosidase and nuclear receptor subfamily 4, group A, member 2 (NR4A2) showed reduced expression. We show that in sporadic PD, parkin, alpha-synuclein and dopamine pathways are co-deregulated. Alpha-synuclein is a member of all three gene regulatory networks. Our analysis results support the view that alpha-synuclein has a central role in the familial as well as the non-familial form of the disease and provide steps towards a pathway definition of PD.
Asunto(s)
Dopamina/metabolismo , Trastornos Parkinsonianos/metabolismo , Trastornos Parkinsonianos/patología , Sustancia Negra/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , alfa-Sinucleína/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Femenino , Regulación de la Expresión Génica/fisiología , Humanos , Masculino , Análisis por Micromatrices/métodos , Persona de Mediana Edad , Modelos Biológicos , Transducción de Señal , Ubiquitina-Proteína Ligasas/genética , alfa-Sinucleína/genéticaRESUMEN
Sporadic Parkinson's disease (PD) is characterized by progressive death of dopaminergic neurons within the substantia nigra. However, pathological cell death within this nucleus is not uniform. In PD, the lateral tier of the substantia nigra (SNl) degenerates earlier and more severely than the more medial nigral component (SNm). The cause of this brain regional vulnerability remains unknown. We have used DNA oligonucleotide microarrays to compare gene expression profiles from the SNl to those of the SNm in both PD and control cases. Genes expressed more highly in the PD SNl included the cell death gene, p53 effector related to PMP22, the tumour necrosis factor (TNF) receptor gene, TNF receptor superfamily, member 21, and the mitochondrial complex I gene, NADH dehydrogenase (ubiquinone) 1beta subcomplex, 3, 12 kDa (NDUFbeta3). Genes that were more highly expressed in PD SNm included the dopamine cell signalling gene, cyclic adenosine monophosphate-regulated phosphoprotein, 21 kDa, the activated macrophage gene, stabilin 1, and two glutathione peroxidase (GPX) genes, GPX1 and GPX3. Thus, there is increased expression of genes encoding pro-inflammatory cytokines and subunits of the mitochondrial electron transport chain, and there is a decreased expression of several glutathione-related genes in the SNl suggesting a molecular basis for pathoclisis. Importantly, some of the genes that are differentially regulated in the SNl are known to be expressed highly or predominantely in glial cells. These findings support the view that glial cells can be primarily affected in PD emphasizing the importance of using a whole tissue approach when investigating degenerative CNS disease.
Asunto(s)
Análisis de Secuencia por Matrices de Oligonucleótidos , Enfermedad de Parkinson/genética , Enfermedad de Parkinson/fisiopatología , ARN Mensajero/genética , Sustancia Negra/fisiopatología , Citocinas/genética , Regulación de la Expresión Génica , Humanos , Inflamación/genética , Mitocondrias/genética , Mitocondrias/fisiología , Proteínas de la Mielina/genética , Proteínas del Tejido Nervioso/genéticaRESUMEN
There is growing evidence that dysfunction of the mitochondrial respiratory chain and failure of the cellular protein degradation machinery, specifically the ubiquitin-proteasome system, play an important role in the pathogenesis of Parkinson's disease. We now show that the corresponding pathways of these two systems are linked at the transcriptomic level in Parkinsonian substantia nigra. We examined gene expression in medial and lateral substantia nigra (SN) as well as in frontal cortex using whole genome DNA oligonucleotide microarrays. In this study, we use a hypothesis-driven approach in analysing microarray data to describe the expression of mitochondrial and ubiquitin-proteasomal system (UPS) genes in Parkinson's disease (PD). Although a number of genes showed up-regulation, we found an overall decrease in expression affecting the majority of mitochondrial and UPS sequences. The down-regulated genes include genes that encode subunits of complex I and the Parkinson's-disease-linked UCHL1. The observed changes in expression were very similar for both medial and lateral SN and also affected the PD cerebral cortex. As revealed by "gene shaving" clustering analysis, there was a very significant correlation between the transcriptomic profiles of both systems including in control brains. Therefore, the mitochondria and the proteasome form a higher-order gene regulatory network that is severely perturbed in Parkinson's disease. Our quantitative results also suggest that Parkinson's disease is a disease of more than one cell class, i.e. that it goes beyond the catecholaminergic neuron and involves glia as well.
Asunto(s)
Encéfalo/metabolismo , Perfilación de la Expresión Génica , Mitocondrias/metabolismo , Enfermedad de Parkinson/metabolismo , Complejo de la Endopetidasa Proteasomal/metabolismo , Estudios de Casos y Controles , Análisis por Conglomerados , Regulación hacia Abajo , Análisis de Secuencia por Matrices de Oligonucleótidos , Sustancia Negra/metabolismo , Ubiquitina/metabolismo , Regulación hacia ArribaRESUMEN
We have used brain tissue from clinically well-documented and neuropathologically confirmed cases of sporadic Parkinson's disease to establish the transcriptomic expression profile of the medial and lateral substantia nigra. In addition, the superior frontal cortex was analyzed in a subset of the same cases. DNA oligonucleotide microarrays were employed, which provide whole human genome coverage. A total of 570 genes were found to be differentially regulated at a high level of significance. A large number of differentially regulated expressed sequence tags were also identified. Levels of mRNA sequences encoded by genes of key interest were validated by means of quantitative real-time polymerase chain reaction (PCR). Comparing three different normalization procedures, results based on the recently published GeneChip Robust Multi Array algorithm were found to be the most accurate predictor of real-time PCR results. Several new candidate genes which map to PARK loci are reported. In addition, the DNAJ family of chaperones is discussed in the context of Parkinson's disease pathogenesis.
Asunto(s)
Perfilación de la Expresión Génica , Genoma Humano , Enfermedad de Parkinson/genética , Sustancia Negra , Anciano , Anciano de 80 o más Años , Femenino , Regulación de la Expresión Génica , Humanos , Masculino , Persona de Mediana Edad , Datos de Secuencia Molecular , Análisis de Secuencia por Matrices de Oligonucleótidos , Reproducibilidad de los Resultados , Sustancia Negra/anatomía & histología , Sustancia Negra/fisiologíaRESUMEN
The cell culture model utilized in this study represents one of the most widely used paradigms of microglia in vitro. After 14 days, microglia harvested from the neonatal rat brain are considered 'mature'. However, it is clear that this represents a somewhat arbitrary definition. In this paper, we provide a transcriptome definition of such microglial cells. More than 7,000 known genes and 1,000 expressed sequence tag clusters were analysed. 'Microglia genes' were defined as sequences consistently expressed in all microglia samples tested. Accordingly, 388 genes were identified as microglia genes. Another 1,440 sequences were detected in a subset of the cultures. Genes consistently expressed by microglia included genes known to be involved in the cellular immune response, brain tissue surveillance, microglial migration as well as proliferation. The expression profile reported here provides a baseline against which changes of microglia in vitro can be examined. Importantly, expression profiling of normal microglia will help to provide the presently purely operational definition of 'microglial activation' with a molecular biological correlate. Furthermore, the data reported here add to our understanding of microglia biology and allow projections as to what functions microglia may exert in vivo, as well as in vitro.
Asunto(s)
Técnicas de Cultivo de Célula , Perfilación de la Expresión Génica , Microglía/citología , Microglía/fisiología , Animales , Animales Recién Nacidos , Células Cultivadas , Etiquetas de Secuencia Expresada , Datos de Secuencia Molecular , Familia de Multigenes , Análisis de Secuencia por Matrices de Oligonucleótidos , Ratas , Ratas Endogámicas LewRESUMEN
This study provides an expression signature of interferon-gamma (IFN-gamma)-activated microglia. Microglia are macrophage precursor cells residing in the brain and spinal cord. The microglial phenotype is highly plastic and changes in response to numerous pathological stimuli. IFN-gamma has been established as a strong immunological activator of microglial cells both in vitro and in vivo. Affymetrix RG_U34A microarrays were used to determine the effect of IFN-gamma stimulation on migroglia cells isolated from newborn Lewis rat brains. More than 8,000 gene sequences were examined, i.e., 7,000 known genes and 1,000 expressed sequence tag (EST) clusters. Under baseline conditions, microglia expressed 326 of 8,000 genes examined (approximately 4% of all genes, 182 known and 144 ESTs). Transcription of only 34 of 7,000 known genes and 8 of 1,000 ESTs was induced by IFN-gamma stimulation. The majority of the newly expressed genes encode pro-inflammatory cytokines and components of the MHC-mediated antigen presentation pathway. The expression of 60 of 182 identified genes and of 9 of 144 ESTs was increased by IFN-gamma, whereas 29 of 182 known genes and 7 of 144 ESTs were down-regulated or undetectable in IFN-gamma-stimulated cultures. Overall, the activating effect of IFN-gamma on the microglial transcriptome showed restriction to pathways involved in antigen presentation, protein degradation, actin binding, cell adhesion, apoptosis, and cell signaling. In comparison, down-regulatory effects of IFN-gamma stimulation appeared to be confined to pathways of growth regulation, remodeling of the extracellular matrix, lipid metabolism, and lysosomal processing. In addition, transcriptomic profiling revealed previously unknown microglial genes that were de novo expressed, such as calponin 3, or indicated differential regulatory responses, such as down-regulation of cathepsins that are up-regulated in response to other microglia stimulators.
Asunto(s)
Antineoplásicos/farmacología , Interferón gamma/farmacología , Microglía/fisiología , Análisis de Secuencia por Matrices de Oligonucleótidos , Transcripción Genética/efectos de los fármacos , Animales , Células Cultivadas , Regulación hacia Abajo/efectos de los fármacos , Expresión Génica/efectos de los fármacos , Expresión Génica/fisiología , Microglía/citología , Microglía/efectos de los fármacos , Análisis de Secuencia por Matrices de Oligonucleótidos/normas , Ratas , Ratas Endogámicas Lew , Reproducibilidad de los Resultados , Transcripción Genética/fisiología , Regulación hacia Arriba/efectos de los fármacosRESUMEN
The discovery that missense mutations in the alpha-synuclein gene represent a rare genetic cause of Parkinson's disease (PD) has had significant impact on the development of research into neurodegenerative disorders. It is becoming increasingly clear that alpha-synuclein plays a central role in the pathological process, which causes Lewy body formation and neurodegeneration in PD. Importantly, there is evidence to suggest that mutated alpha-synuclein is toxic to both nerve cells and glia. However, the regulation and function of wild-type alpha-synuclein are as yet ill defined. Using the facial nerve axotomy model, we have addressed the question whether the expression of alpha-synuclein in nerve cells may change in response to injury. We were particularly interested in testing the hypothesis that the severity of neuronal injury had an effect on alpha-synuclein metabolism. Facial nerve cut and crush, respectively, were performed in adult rats where normal facial motoneurones do not express alpha-synuclein. Following axotomy, a subset of facial motoneurones newly expressed high levels of alpha-synuclein immunoreactivity in their cell body and, occasionally, their nucleus. Significantly more nerve cells were labelled following facial nerve transection than following facial nerve crush. Confocal microscopy revealed a granular pattern of alpha-synuclein aggregation in degenerating nerve cells. Interestingly, the observed cell death phenotype was clearly non-apoptotic and developed over days or weeks rather than hours. Thus, axotomy of adult rat facial motoneurones triggers de novo expression of alpha-synuclein and this expression is associated with a non-apoptotic, slow form a neurodegeneration. In addition, the extent of alpha-synuclein expression is related to the severity of neuronal injury.
Asunto(s)
Nervio Facial/metabolismo , Neuronas Motoras/metabolismo , Degeneración Nerviosa/metabolismo , Proteínas del Tejido Nervioso/biosíntesis , Animales , Apoptosis/fisiología , Axotomía/métodos , Nervio Facial/patología , Neuronas Motoras/patología , Compresión Nerviosa/métodos , Degeneración Nerviosa/patología , Ratas , Ratas Endogámicas Lew , Sinucleínas , alfa-SinucleínaRESUMEN
We have exploited a procedure to identify when hydrogen bonds (H-bonds) form under two-state folding conditions using equilibrium and kinetic deuterium/hydrogen amide isotope effects. Deuteration decreases the stability of equine cytochrome c and the dimeric and crosslinked versions of the GCN4-p1 coiled coil by approximately 0. 5 kcal mol-1. For all three systems, the decrease in equilibrium stability is reflected by a decrease in refolding rates and a near equivalent increase in unfolding rates. This apportionment indicates that approximately 50% of the native H-bonds are formed in the transition state of these helical proteins. In contrast, an alpha/beta protein, mammalian ubiquitin, exhibits a small isotope effect only on unfolding rates, suggesting its folding pathway may be different. These four proteins recapitulate the general trend that approximately 50% of the surface buried in the native state is buried in the transition state, leading to the hypothesis that H-bond formation in the transition state is cooperative, with alpha-helical proteins forming a number of H-bonds proportional to the amount of surface buried in the transition state.
Asunto(s)
Amidas/metabolismo , Proteínas de Unión al ADN , Deuterio/metabolismo , Enlace de Hidrógeno , Pliegue de Proteína , Proteínas/química , Proteínas de Saccharomyces cerevisiae , Amidas/química , Sustitución de Aminoácidos , Animales , Grupo Citocromo c/química , Grupo Citocromo c/metabolismo , Dimerización , Disulfuros/metabolismo , Estabilidad de Enzimas , Proteínas Fúngicas/química , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Humanos , Isomerismo , Cinética , Modelos Moleculares , Prolina/química , Prolina/metabolismo , Desnaturalización Proteica , Proteínas Quinasas/química , Proteínas Quinasas/genética , Proteínas Quinasas/metabolismo , Estructura Secundaria de Proteína , Proteínas/metabolismo , Solventes , Termodinámica , Ubiquitinas/química , Ubiquitinas/metabolismo , Agua/química , Agua/metabolismoRESUMEN
We have investigated the folding behavior of dimeric and covalently crosslinked versions of the 33-residue alpha-helical GCN4-p1 coiled coil derived from the leucine zipper region of the transcriptional activator GCN4. The effects of multisite substitutions indicate that folding occurs along multiple routes with nucleation sites located throughout the protein. The similarity in activation energies of the different routes together with an analysis of intrinsic helical propensities indicate that minimal helix is present before a productive collision of the two chains. However, approximately one-third to one-half of the total helical structure is formed in the postcollision transition state ensemble. For the crosslinked, monomeric version, folding occurs along a single robust pathway. Here, the region nearest the crosslink, with the least helical propensity, is structured in the transition state whereas the region farthest from the tether, with the most propensity, is completely unstructured. Hence, the existence of transition state heterogeneity and the selection of folding routes critically depend on chain topology.
Asunto(s)
Proteínas de Unión al ADN , Proteínas Fúngicas/química , Proteínas Quinasas/química , Proteínas de Saccharomyces cerevisiae , Secuencia de Aminoácidos , Núcleo Celular/química , Dicroismo Circular , Relación Dosis-Respuesta a Droga , Guanidina/farmacología , Modelos Biológicos , Datos de Secuencia Molecular , Mutagénesis , Biosíntesis de Péptidos , Pliegue de Proteína , Estructura Secundaria de Proteína , TermodinámicaRESUMEN
A new technique for detecting spectral spin diffusion in solids under MAS NMR conditions that is particularly well suited for accurately measuring cross-relaxation from minor spectral components is presented. The pulse sequence, SINK (Saturation Inter-Nuclear Kinetics), selectively saturates the magnetization of a minor spectral component with a series of rotor-synchronized DANTE pulse trains and monitors spin diffusion to other peaks with a non-selective 90 degrees pulse. We have used SINK with 19F MAS NMR on samples of calcium fluorapatite doped with Sb3+ to measure spin diffusion between a weak peak at 68.6 ppm due to fluoride ions associated with Sb3+ and other peaks in the spectrum. The SINK experiment clearly demonstrates that spin diffusion from the former peak to the main resonance of fluorapatite at 64.0 ppm is faster than spin diffusion to a second antimony-related peak at 65.6 ppm. These results strengthen our previous conclusion that antimony(III) occupies a phosphate site in the apatite lattice, with an SbO3(3-) group replacing a PO4(3-) group. The SINK experiment also enables the detection of a "hidden" peak at approximately 62.9 ppm that is otherwise obscured by the intense main peak at 64.0 ppm.
Asunto(s)
Apatitas/química , Espectroscopía de Resonancia Magnética/métodos , Antimonio/química , Flúor/química , Modelos Químicos , Estructura MolecularRESUMEN
Subcloning of a clone of the 120-bp family of rye, pSc119, has produced two extremely useful probes. pSc119.1 assays rye-specific dispersed repetitive sequence families. It is present on all seven rye chromosomes and hybridizes to the entire length of each chromosome, with the exception of some telomeres and the nucleolar organiser region. pSc119.2, in contrast, hybridizes predominantly to the telomeric regions of rye chromosomes, with some interstitial sites. Unlike pSc119.1, it assays similar repetitive sequence families in both wheat and rye chromosomes.
Asunto(s)
Cromosomas , ADN/genética , Secuencias Repetitivas de Ácidos Nucleicos , Secale/genética , Triticum/genética , Células Clonales , Sondas de ADN , Cariotipificación , Datos de Secuencia Molecular , Hibridación de Ácido NucleicoRESUMEN
An analysis of four species from the genus Secale, including the study of different accessions, has shown that the properties of DNA clones of monomer units from three repeated sequence loci, namely, Ter, Nor, and 5S DNA, proved to be representative of the entire loci from which they were isolated. This finding in Secale species, including the discovery of a new locus for 5S DNA on chromosome 5R, has been used to interpret information on the Ter, Nor, and 5S DNA loci from 15 species in the Triticeae complex. The evolutionary relationship among species suggested by the DNA sequence data has shown many consistencies with a number of other characters such as those used in classical systematics, as well as geographical distribution data and isozyme and chromosome-pairing studies. Apparent inconsistencies such as a close relationship between the R and P genomes at the Ter loci are interpreted in terms of amplification-deletion phenomena known to occur at repetitive sequence loci. In addition, this study included species endemic to Australia and thus provided a broad time span in which to consider some features of repeated sequence family evolution, such as the conservation of certain parts of 5S DNA spacer regions.