Nucleic acid encoding CLK2 protein kinases

Details Nucleic acid encoding CLK2 protein kinases Nucleic acid encoding CLK2 protein kinases

2001-07-17

2004-09-28

Spector, Lorraine (Department: 1646)

Chemistry: molecular biology and microbiology

Animal cell, per se ; composition thereof; process of...

C435S069100, C435S194000, C435S320100, C435S252300, C435S254110, C536S023100, C536S023500, C530S300000, C530S350000

Reexamination Certificate

active

06797513

ABSTRACT:

INTRODUCTION
The present invention relates to novel CDC2 like protein kinases (CLK protein kinases). These protein kinases phosphorylate proteins rich in serine and arginine.
BACKGROUND OF THE INVENTION
The following description of the background of the invention is provided to aid in understanding the invention, but is not admitted to be or describe prior art to the invention.
Cellular signal transduction is a fundamental mechanism whereby extracellular stimuli are relayed to the interior of cells and subsequently regulate diverse cellular processes. One of the key biochemical mechanisms of signal transduction involves the reversible phosphorylation of proteins. Phosphorylation of polypeptides regulates the activity of mature proteins by altering their structure and function. Phosphate most often resides on the hydroxyl moiety (—OH) of serine, threonine, or tyrosine amino acids in proteins. Enzymes that mediate phosphorylation of cellular effectors fall into two classes. While protein phosphatases hydrolyze phosphate moieties from phosphoryl protein substrates, protein kinases transfer a phosphate moiety from adenosine triphosphate to protein substrates. The converse functions of protein kinases and protein phosphatases balance and regulate the flow of signals in signal transduction processes.
Protein kinases and protein phosphatases are typically divided into two groups: receptor and non-receptor type proteins. Receptor protein kinases are comprised of an extracellular domain, a membrane spanning region, and a catalytic domain.
Protein kinases and protein phosphatases are divided further into three classes based upon the amino acids they act upon. Some catalyze the addition or hydrolysis of phosphate on serine or threonine only, some catalyze the addition or hydrolysis of phosphate on tyrosine only, and some catalyze the addition or hydrolysis of phosphate on serine, threonine, and tyrosine.
Membrane association is an important feature of signal transduction. Protein kinases propagate extracellular signals to the inside of the cell by attracting other signaling molecules to the membrane. Schlessinger and Ullrich, 1992
, Neuron
9:383-391. For instance, many receptor protein kinases bind an extracellular ligand, dimerize, and cross phosphorylate one another. These phosphate moieties subsequently attract other proteins necessary for propagating the signal within the cell. The molecules that signal downstream of the receptor protein kinases are often nonreceptor protein kinases which propagate and amplify the extracellular signal.
A class of non-receptor protein kinases are implicated in regulating RNA splicing. Fu, 1995
RNA
1:663-680; Staknis and Reed, 1994
, Mol. Cell. Biol.
14:7670-7682. These protein kinases phosphorylate polypeptides rich in serine and arginine (SR proteins). SR proteins are characterized as containing at least one amino-terminal RNA recognition motif and a basic carboxyterminal domain rich in serine and arginine residues, often arranged in tandem repeats. Zahler et al., 1992
, Genes Dev
6:837-847. Experimental evidence supports the idea that the SR domain is involved in protein—protein interactions (Kohtz et al., 1994
, Nature
368:119-124) as well as protein-RNA interactions (Harada et al., 1996
, Nature
380:175-179), and may contribute to a localization signal directing proteins to nuclear speckles. Hedley et al., 1995
, Proc. Natl. Acad. Sci. USA
92:11524-11528.
A recent report demonstrated mCLK1, a CDC2 like kinase, interacts with ASF/SF2, SRp20 and hnRNP proteins in a yeast two hybrid system. Because hnRNP-K binds to the protooncogene p95
vav
, mCLK1 could be implicated in transmitting signals that regulate the expression of the protooncogenes myc and fos in hematopoietic cells. Furthermore, it was demonstrated, that mCLK1 could phosphorylate ASF/SF2 in vitro, suggesting, that SR containing proteins are the natural substrates of mCLK1. Colwill et al., 1996
, EMBO J.
15:265-275.
mCLK1 is a dual specificity protein kinase originally isolated in mouse expression libraries (Ben-David et al., 1991
, EMBO J.
10:317-325; Howell et al., 1991
, Mol. Cell. Biol.
11:568-572) and human (hCLK1, hCLK2, hCLK3), plant (AFC1, AFC2, AFC3) and fly (DOA) CLK protein kinases have since been identified. Johnson and Smith, 1991
, J. Biol. Chem.
266:3402-3407; Hanes et al., 1994
, J. Mol. Biol.
244:665-672; Bender and Fink, 1994
, Proc. Natl. Acad. Sci. USA
91:12105-12109; Yun et al., 1994
, Genes. Dev.
8:1160-1173. The amino terminal domain of these proteins is rich in serine and arginine, whereas the catalytic domain can be most similar to CDC2, a serine/threonine protein kinase. Ben-David et al., 1991
, EMBO J.
10:317-325.
Both mCLK1 and the Drosophila homologue, DOA, regulate RNA splicing events. Each of these have two alternatively spliced products coding for either the full-length catalytically active protein or a truncated protein lacking the catalytic domain. Yun et al., 1994
, Genes. Dev.
8:1160-1173; Duncan et al., 1995
, J. Biol. Chem.
270:21524-21531. Identical splice forms were also found in human CLK protein kinases. Hanes et al., 1994
, J. Mol. Biol.
244:665-672. The ratio of these splice products appears to be developmentally regulated in Drosophila (Yun et al., 1994
, Genes. Dev.
8:1160-1173), and in a tissue and cell type specific manner in mammals. Hanes et al., 1994
, J. Mol. Biol.
244:665-672; Duncan et al., 1995
, J. Biol. Chem.
270:21524-21531. In addition, the expression of several other, larger transcripts, are observed to be differentially regulated and are shown to represent partially spliced products. Duncan et al., 1995
, J. Biol. Chem.
270:21524-21531.
SUMMARY OF THE INVENTION
The present invention is based in part upon the isolation and characterization of nucleic acid molecules encoding CLK serine/threonine kinases designated mCLK2, mCLK3, and mCLK4. CLK serine/threonine kinases regulate RNA splicing in cells and some are highly expressed in cancer cells as well as testis. Various mCLK2, mCLK3, and mCLK4 related molecules and compounds can now be designed as treatments of cancers or as contraceptives to reproduction in male organisms.
The present invention is based in part upon nucleic acid molecules encoding novel mCLK2, mCLK3, and mCLK4 polypeptides, nucleic acid molecules encoding portions of their amino acid sequences, nucleic acid vectors harboring such nucleic acid molecules, cells containing such nucleic acid vectors, purified polypeptides encoded by such nucleic acid molecules, and antibodies to such polypeptides, and methods of identifying compounds that bind mCLK2, mCLK3, and mCLK4 or abrogate their interactions with natural binding partners. Also disclosed are methods for diagnosing and treating specific abnormal conditions in an organism with mCLK2, mCLK3, and mCLK4 related molecules or compounds. The nucleic acid molecules, nucleic acid vectors, recombinant cells, polypeptides, and antibodies may be produced using well known and standard techniques used currently in the art.
Thus in a first aspect, the invention features isolated, enriched, or purified nucleic acid molecules encoding a novel mCLK2, mCLK3, or mCLK4 polypeptide.
The term “isolated”, in reference to nucleic acid molecules, indicates that a naturally occurring sequence has been removed from its normal cellular environment. The isolated nucleic acid of the present invention is unique in the sense that it is not found in a pure or separated state in nature. Use of the term “isolated” indicates that a naturally occurring sequence has been removed from its normal cellular (i.e., chromosomal) environment. Thus, the sequence may be in a cell-free solution or placed in a different cellular environment. The term does not imply that the sequence is the only nucleotide chain present, but that it is essentially free (about 90-95% pure at least) of non-nucleotide material naturally associated with it, and thus is distinguished from isolated chromosomes.
The term “enriched”, in reference to nucleic acid molecules, means that the specific DNA or RNA sequence

Affiliated with

Also associated with

Rating

Nucleic acid encoding CLK2 protein kinases does not yet have a rating. At this time, there are no reviews or comments for this patent.

If you have personal experience with Nucleic acid encoding CLK2 protein kinases, we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Nucleic acid encoding CLK2 protein kinases will most certainly appreciate the feedback.

Rate now

Comments { 0 }

Profile ID: LFUS-PAI-O-3245975