The number and intricacy of processes necessary for the organisation and sustenance of life demands a fair degree of genetic and genomic complexity. Recent evaluations of the human genome estimate our complement of genes in the order of >30,000 represented in over 2.91 billion base pairs of DNA. Given that most cells in the body are diploid, this makes for a great deal of material that has to be carefully ordered in the nucleus, such that precise activity of these genes remains possible. Achieving this level of organisation is accomplished through interactions between the DNA and a number of proteins.

The nuclear matrix is a network of proteins that provides a structural framework for organising chromatin, while facilitating transcription and replication. Until recently attachment to the nuclear matrix was regarded as static, functionally segregating chromatin into loop domains. At present, a more dynamic model is being considered. In this model, cellular function and state-dependent interactions with the nuclear matrix provide a means by which segments of chromatin can acquire or maintain the open, potentiated conformation necessary for transcription. The human protamine gene cluster, and more recently, the human beta-gloin gene cluser have been used as model systems to address how matrix association may mediate the spatial and temporal patterns of gene expression.

Our work has also involved the development of a computer algorithm use to predict to predict putative sites of nuclear matrix association called MAR-wiz. This program uses sequence data as an input and plots the matrix association pontential along the query sequence based on several motifs which have been shown to associate with the nuclear matrix.

This plot shows the matrix binding potential of a fragment of chromosome 16 encompassing the human protamine locus (GenBank accession number U15422). Predicted regions of high binding potential (>0.75) correspond to those experimentally verified (red boxes: haploid-specific protamine MARs; blue box: somatic-SOCS1 MAR).

MARs and the regulation of the b-globin locus. (a) b-globin non-expressing cells. The b-globin locus is comprised of the epsilon>G-gamma>A-gamma>delta>beta-globin genes. These genes are expressed in a time- and spatial-dependent manner. Attachment is indicated by outlined segments adjacent to the nuclear matrix. Attachment of the LCR/5'HS region of the human globin locus in non-erythroid cell lines is proposed to play a role in quenching expression. Two additional regions are tethered to the nuclear matrix between the LCR and b-globin gene to spatially separate and silence the locus. (b) Cells permissive for b-globin locus expression. In response to a maturation signal, the previously tethered regions between the LCR/5'HS and the b-globin gene are released from the nuclear matrix and the chromatin domain is remodeled. With new nuclear matrix tethers, a chromatin conformation is then established that facilitates looping of the active genes back to the LCR/5'HS region. [From Ostermeier et al. 2003 go to publications...]

top image: HeLa cell nuclei; histones extracted with 25mM 3.5-diiodosalicylic acid (lithium salt); time course: 0-15min. Nuclei are stained with DAPI to visualise DNA. Loops of matrix independent DNA become clear over time while matrix-associated DNA remains bound to remaining nuclear matrix elements. Extracted nuclei are shown at >100X magnification, with a solarising filter.


TEMicrograph of mouse fibroblast nuclear matrix[left] and surrounding cytoplasm . Fibroblasts were detergent extracted and DNaseI treated.
(Capco et al. 1982)


  references

Kramer, J.A., Singh, G.B. and Krawetz, S.A. (1996) Computer assisted search for sites of nuclear matrix attachment. Genomics 33:305-308. [PubMed]

Kramer, J.A. and Krawetz, S.A. (1996) Nuclear matrix interactions within the sperm genome. The Journal of Biological Chemistry 271:11619-11622. [PubMed]

Singh, G.B., Kramer, J.A. and Krawetz, S.A. (1997) Mathematical model to predict regions of chromatin attachment to the nuclear matrix. Nucleic Acids Research 25:1419-1425. [PubMed]

Kramer, J.A. and Krawetz, S.A. (1997) A PCR-based assay to determine nuclear matrix association. BioTechniques 22:826-828. [PubMed]

Kramer, J.A. and Krawetz, S.A. (1997) Determining the potentiative state of a chromatin domain. BioTechniques 22:880-882. [PubMed]

Kramer, J.A., Zhang, S., Yaron, Y., Zhao, Y. and Krawetz, S.A. (1997) Genetic testing for male infertility: a postulated role for mutations in sperm nuclear matrix attachment regions. Genetic Testing 1:125-129. [PubMed]

Singh, G.B. and Krawetz, S.A. (2000) Data mining algorithm for discovering matrix association regions (MARs) Proceedings of Spie, Data Mining and Knowledge Discovery: Theory, Tools, and Technology II, 4057:330-341.

Schmid, C., Heng, H.H.Q., Rubin, C., Ye, C.J. and Krawetz S.A. (2001) Sperm nuclear matrix association of the PRM1>PRM2>TNP2 domain is independent of Alu Methylation, Human Molecular Reproduction 7:903-911. [PubMed]

Ostermeier, G.C., Liu, Z., Martins, R.P., Bharadwaj R.R., Ellis J., Draghici, S.,and Krawetz, S.A. (2003) Nuclear matrix association of the human beta-globin locus utilizing a novel approach to quantitative real-time PCR. Nucleic Acids Research. 31:3257-3266.  [PubMed]

 


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