Telokin

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Telokin domain
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Telokin
Telokin.png
Crystallographic structure of muscle myosin light chain kinase from turkey gizzard. [1]
Identifiers
Symbol MYLK
NCBI gene 4638
HGNC 7590
OMIM 600922
RefSeq NM_053025
UniProt Q5MYA0
Other data
EC number 2.7.11.18
Locus Chr. 3 q21

Telokin (also known as kinase-related protein or KRP) is an abundant protein found in smooth-muscle. It is identical to the C-terminus of myosin light-chain kinase. Telokin may play a role in the stabilization of unphosphorylated smooth-muscle myosin filaments. [2] [3] Because of its origin as the C-terminal end of smooth muscle myosin light chain kinase, it is called "telokin" (from a combination of the Greek telos, "end" and kinase). [4]

Contents

Nomenclature and classification

Telokin's systematic name is ATP:[myosin light chain] O-phosphotransferase and its recommended name is myosin-light-chain kinase. (EC 2.7.11.18).

The gene MYLK, a muscle member of the immunoglobulin gene superfamily, encodes myosin light chain kinase which is a calcium/calmodulin dependent enzyme. Four transcript variants that produce four isoforms of the calcium/calmodulin dependent enzyme have been identified as well as two transcripts that produce two isoforms of telokin. The two transcripts that produce the two telokin isoforms are the following:

Isoform 7
This variant encodes the shorter isoform of kinase related protein, telokin. The first exon corresponds to intron 30 and the remainder of the transcript corresponds to the last two exons of the gene. It is shorter than variant 8 by one codon at the splicing junction between the first two exons. It is made by 153 aa. NCBI Reference Sequence: NP_444259.1. It comes from the Homo sapiens myosin light chain kinase (MYLK), transcript variant 7, mRNA, whose length is 2676 bp. NCBI Reference Sequence: NM_053031.2.
Isoform 8
This variant encodes the longer isoform of kinase related protein, telokin. It is longer than variant 7 by one codon at the splicing junction between the first two exons. It is composed of 154 aa. NCBI Reference Sequence: NP_444260.1. It comes from the Homo sapiens myosin light chain kinase (MYLK), transcript variant 8, mRNA, whose length is 2679 bp. NCBI Reference Sequence: NM_053032.2.

Catalytic activity and other functional data

Telokin catalyzes the following reaction:

It requires Ca2+ and calmodulin for activity. The 20-kDa light chain from smooth muscle myosin is phosphorylated more rapidly than any other acceptor, but light chains from other myosins and myosin itself can act as acceptors, but more slowly. [5]

The Km values of homo sapiens telokin is 0.018 mM at 23–25 °C and pH = 7.5. This enzyme has a pH optimum of 7.4 and temperature optimum of 30 °C. [6]

Telokin is an acidic protein with a PI value of 4-5 and 17-kDa with an amino acid sequence that is identical to the C terminus of the 130-kDa myosin light chain kinase (MLCK), although it is expressed as a separate protein and produced by an alternate promoter of the MLCK gene. Telokin is transcribed from a second promoter, located within an intron, in the 3' region of the MLCK gene. [7] And that is why the concentration of telokin (at least 15 μM) is higher than MLCK concentration.

Telokin has been shown to bind to unphosphorylated myosin filaments and to stimulate myosin mini-filament assembly in vitro. The major mechanism for initiating smooth muscle (SM)2 contraction is the rise in Ca2+ concentration resulting in an increase in 20-kDa myosin regulatory light chain (MLC20) phosphorylation at Ser-19.

Structure

Primary structure
Telokin is an intracellular protein and, as such, does not contain the disulfide linkage between beta-strands B and F normally observed in the immunoglobulin constant domains. It does, however, contain two cysteine amino acid residues (Cys63 and Cys115) that are situated at structurally identical positions to those forming the disulfide linkage in the immunoglobulin constant domain.
Secondary structure
Telokin contains 154 amino acid residues, 103 of which were visible in the electron density map. Telokin and the C-terminal domain of MLCK show amino acid sequence similarity to several quite unrelated muscle proteins such as titin or C-protein. [2] The overall molecular fold of telokin consists of seven strands of antiparallel beta-pleated sheet that wrap around to form a barrel. There is also an extended tail of eight amino acid residues at the N terminus that does not participate in beta-sheet formation. The beta-barrel can be simply envisioned as two layers of beta-sheet, nearly parallel to one another, with one layer containing four and the other three beta-strands.
Domains
Telokin has a particular domain called Ig-like I-type (Immunoglobulin like intermediate type) with a length of 92 residues between 42 and 133. At the beginning it was thought that this domain was Ig- like C2-type but some studies determined that its structure shares characteristics from V-set and C2-set and that is why the a I-type was invented. These kind of domains mediates T-cell adhesion via its ectodomain, and signal transduction.

Tissue distribution

KRP presence in different tissues has been assessed by immunoblots using anti-KRP antibodies, and by analyses of its mRNA in Northern blot. [8] [9] [10] KRP is an abundant smooth muscles-pecific protein. So far it has not been detected in non-muscle tissues and striated muscles. [9] Its concentration in gizzard muscle is lo-12-fold higher than that of MLCK and only 2-3-fold less than that of myosin. [8] Vascular muscles have a lower KRP/MLCK ratio. [9] [11]

Telokin is expressed at very high levels in intestinal, urinary, and reproductive tract smooth muscle, at lower levels in vascular smooth muscle, and at undetectable levels in skeletal or cardiac muscle or nonmuscle tissues. Although telokin is strongly activated by myocardin and myocardin is highly expressed in vascular smooth muscle cells, there is relatively little expression of telokin in these cells. This suggests that an inhibitory factor must be attenuating the activity of the telokin promoter in vascular smooth muscle cells. One possible candidate for this inhibitory factor is GATA-6 [12]

The increase in telokin expression correlated with an increase in the expression of several other smooth muscle-restricted proteins, including smooth muscle myosin and alpha-actin. [7]

Accumulates in individuals with asthma (at protein levels). [13] Induced by tumor necrosis factor (TNF). [14] Repressed by androgens (e.g. R1881). [15]

Function

Telokin have two related functions in the C-terminal myosin-binding domain of smooth muscle myosin light chain kinase (MLCK). First, telokin stabilizes myosin filaments in the presence of ATP. Second, telokin can modulate the level of myosin light chain phosphorylation. In this latter role, multiple mechanisms have been suggested. One hypothesis is that light chain phosphorylation is diminished by the direct competition of KRP and MLCK for myosin, resulting in a loss of contraction. [8]

Telokin also inhibits the phosphorylation of myosin filaments while having no effect on phosphorylation of the isolated smooth-muscle myosin regulatory light chain (ReLC). However, when telokin was phosphorylated by MLCK, the telokin-induced inhibition of myosin phosphorylation was removed, which indicates the existence of a telokin-dependent modulatory pathway in smooth-muscle regulation. In this part we must say that the phosphorylation of telokin can be enhanced by the concentration of Ca2+ and calmodulin.

Kinase-related protein (telokin) binds to dephosphorylated smooth myosin near the junction between the rod and the catalytic head region (S-I). This interaction is prevented by MLCK-catalysed phosphorylation of myosin and conversely, the rate of myosin phosphorylation is in turn inhibited by KRP in vitro. [8] As a consequence of this, in vivo KRP might slow down the rate of myosin phosphorylation by myosin light chain kinase (MLCK) and, therefore, tension development. When the intracellular Ca2+ level is decreasing, the KRP can also accelerate muscle relaxation by lowering the ratio of phosphorylated to phosphorylated myosin. KRP is also an important structural regulator of myosin filaments. Smooth muscle myosin, under physiological conditions in vitro, can adapt two relatively and different stable conformations. When the myosin is in the extended conformation, it is active and able to combine with other myosin molecules to form thick filaments which are fundamental for effective contraction. Upon ATP binding, the rod part of unphosphorylated myosin molecule folds into thirds, so that the head –rod junction is brought close to the middle of the rod and stabilized there, presumable by interacting with both the 20 KDa light chains and the neck region. This interaction is prevented by the MLCK-dependent light chain phosphorylation, resulting in the unfolding of myosin monomers and their rapid polymerization into filaments.

The binding of KRP to the neck region folded, ATP-bound dephosphorylated myosin also promotes unfolding and filament formation, thus looking like light chain phosphorylation. This could be a physiologically significant phenomenon considering the high concentration of ATP always present in smooth muscle cells. Thus, Kinase-related protein may have a very important role in relaxed smooth muscle by keeping dephosphorylated myosin in the filamentous state ready for the next rapid contractile response. Experiments aimed at testing this hypothesis indicated that under appropriate conditions a small excess of KRP is enough to form an equimolar complex with smooth muscle myosin and to cause its complete polymerization in the presence of ATP. Experiments where it has been tested this hypothesis, indicated that in appropriated conditions, a small excess of KRP is enough to form an equimolar complex with smooth muscle myosin and in the presence of ATP, cause its complete polymerization. [11]

Pathology

Certain mutations in the MYLK gene are associated with thoracic aortic aneurysms or thoracic aortic dissections. This disease is caused by mutations affecting the gene MYLK. A disease characterized by permanent dilation of the thoracic aorta usually due to degenerative changes in the aortic wall. It is primarily associated with a characteristic histologic appearance known as 'medial necrosis' or 'Erdheim cystic medial necrosis' in which there is degeneration and fragmentation of elastic fibers, loss of smooth muscle cells, and an accumulation of basophilic ground substance. [16] [17]

The effect of hypoxia

In cats, telokin expression varies inversely with artery diameter except for cerebral arteries where no telokin is observed. Telokin and myosin light chain are distributed uniformly throughout small pulmonary arteries however they do not colocalized. During hypoxia, telokin dephosphorylates, and myosin light chain becomes increasingly phosphorylated in small pulmonary arteries smooth muscle cell, whereas in large pulmonary arteries smooth muscle cell there is no change in either telokin or myosin light chain phosphorylation. When large pulmonary arteries smooth muscle cell were exposed to phenylephrine, myosin light chain phosphorylation increased with no change in telokin phosphorylation. In small pulmonary arteries, phosphorylated telokin may help maintain relaxation under unstimulated conditions, whereas in large pulmonary arteries, telokin's function remains undetermined. [18]

Related Research Articles

Smooth muscle Involuntary non-striated muscle

Smooth muscle is an involuntary non-striated muscle, so-called because it has no sarcomeres and therefore no striations. It is divided into two subgroups, single-unit and multiunit smooth muscle. Within single-unit muscle, the whole bundle or sheet of smooth muscle cells contracts as a syncytium.

Protein kinase A

In cell biology, protein kinase A (PKA) is a family of enzymes whose activity is dependent on cellular levels of cyclic AMP (cAMP). PKA is also known as cAMP-dependent protein kinase. PKA has several functions in the cell, including regulation of glycogen, sugar, and lipid metabolism.

Myosin

Myosins are a superfamily of motor proteins best known for their roles in muscle contraction and in a wide range of other motility processes in eukaryotes. They are ATP-dependent and responsible for actin-based motility. The term was originally used to describe a group of similar ATPases found in the cells of both striated muscle tissue and smooth muscle tissue. Following the discovery by Pollard and Korn (1973) of enzymes with myosin-like function in Acanthamoeba castellanii, a global range of divergent myosin genes have been discovered throughout the realm of eukaryotes.

Muscle contraction Activation of tension-generating sites in muscle

Muscle contraction is the activation of tension-generating sites within muscle cells. In physiology, muscle contraction does not necessarily mean muscle shortening because muscle tension can be produced without changes in muscle length, such as when holding a heavy book or a dumbbell at the same position. The termination of muscle contraction is followed by muscle relaxation, which is a return of the muscle fibers to their low tension-generating state.

CAMK, also written as CaMK, is an abbreviation for the Ca2+/calmodulin-dependent protein kinase class of enzymes. CAMKs are activated by increases in the concentration of intracellular calcium ions (Ca2+) and calmodulin. When activated, the enzymes transfer phosphates from ATP to defined serine or threonine residues in other proteins, so they are serine/threonine-specific protein kinases. Activated CAMK is involved in the phosphorylation of transcription factors and therefore, in the regulation of expression of responding genes. CAMK also works to regulate the cell life cycle (i.e. programmed cell death), rearrangement of the cell's cytoskeletal network, and mechanisms involved in the learning and memory of an organism.

Myosin light-chain kinase Class of kinase enzymes

Myosin light-chain kinase also known as MYLK or MLCK is a serine/threonine-specific protein kinase that phosphorylates a specific myosin light chain, namely, the regulatory light chain of myosin II.

Caldesmon

Caldesmon is a protein that in humans is encoded by the CALD1 gene.

ROCK1

ROCK1 is a protein serine/threonine kinase also known as rho-associated, coiled-coil-containing protein kinase 1. Other common names are ROKβ and P160ROCK. ROCK1 is a major downstream effecter of the small GTPase RhoA and is a regulator of the actomyosin cytoskeleton which promotes contractile force generation. ROCK1 plays a role in cancer and in particular cell motility, metastasis, and angiogenesis.

Myosin-light-chain phosphatase

Myosin light-chain phosphatase, more commonly called myosin phosphatase, is an enzyme that dephosphorylates the regulatory light chain of myosin II. This dephosphorylation reaction occurs in smooth muscle tissue and initiates the relaxation process of the muscle cells. Thus, myosin phosphatase undoes the muscle contraction process initiated by myosin light-chain kinase. The enzyme is composed of three subunits: the catalytic region, the myosin binding subunit (MYPT1), and a third subunit (M20) of unknown function. The catalytic region uses two manganese ions as catalysts to dephosphorylate the light-chains on myosin, which causes a conformational change in the myosin and relaxes the muscle. The enzyme is highly conserved and is found in all organisms’ smooth muscle tissue. While it is known that myosin phosphatase is regulated by rho-associated protein kinases, there is current debate about whether other molecules, such as arachidonic acid and cAMP, also regulate the enzyme.

Myosin light chain Small polypeptide subunit of myosin

A myosin light chain is a light chain of myosin. Myosin light chains were discovered by Chinese biochemist Cao Tianqin when he was a graduate student at the University of Cambridge in England.

CAMK2G

Calcium/calmodulin-dependent protein kinase type II gamma chain is an enzyme that in humans is encoded by the CAMK2G gene.

MYH10

Myosin-10 also known as myosin heavy chain 10 or non-muscle myosin IIB (NM-IIB) is a protein that in humans is encoded by the MYH10 gene. Non-muscle myosins are expressed in a wide variety of tissues, but NM-IIB is the only non-muscle myosin II isoform expressed in cardiac muscle, where it localizes to adherens junctions within intercalated discs. NM-IIB is essential for normal development of cardiac muscle and for integrity of intercalated discs. Mutations in MYH10 have been identified in patients with left atrial enlargement.

MYL2

Myosin regulatory light chain 2, ventricular/cardiac muscle isoform (MLC-2) also known as the regulatory light chain of myosin (RLC) is a protein that in humans is encoded by the MYL2 gene. This cardiac ventricular RLC isoform is distinct from that expressed in skeletal muscle (MYLPF), smooth muscle (MYL12B) and cardiac atrial muscle (MYL7).

MYLK Gene of the immunoglobulin superfamily

Myosin light chain kinase, smooth muscle also known as kinase-related protein (KRP) or telokin is an enzyme that in humans is encoded by the MYLK gene.

MYL7

Atrial Light Chain-2 (ALC-2) also known as Myosin regulatory light chain 2, atrial isoform (MLC2a) is a protein that in humans is encoded by the MYL7 gene. ALC-2 expression is restricted to cardiac muscle atria in healthy individuals, where it functions to modulate cardiac development and contractility. In human diseases, including hypertrophic cardiomyopathy, dilated cardiomyopathy, ischemic cardiomyopathy and others, ALC-2 expression is altered.

MYLK3

Myosin light chain kinase 3 also known as MYLK3, is an enzyme which in humans is encoded by the MYLK3 gene.

MYLK2

Myosin light chain kinase 2 also known as MYLK2 is an enzyme which in humans is encoded by the MYLK2 gene.

MYLK4

Myosin light chain kinase 4 also known as MYLK4 is an enzyme which in humans is encoded by the MYLK2 gene. MYLK4 is a member of the myosin light-chain kinase family of serine/threonine-specific protein kinases that phosphorylate the regulatory light chain of myosin II.

Rho-associated protein kinase

Rho-associated protein kinase (ROCK) is a kinase belonging to the AGC family of serine-threonine kinases. It is involved mainly in regulating the shape and movement of cells by acting on the cytoskeleton.

Calponin 1

Calponin 1 is a basic smooth muscle protein that in humans is encoded by the CNN1 gene.

References

  1. ↑ PDB: 1FHG ​; Holden HM, Ito M, Hartshorne DJ, Rayment I (October 1992). "X-ray structure determination of telokin, the C-terminal domain of myosin light chain kinase, at 2.8 A resolution". J. Mol. Biol. 227 (3): 840–51. doi:10.1016/0022-2836(92)90226-A. PMID   1404391.
  2. 1 2 Sobieszek A, Andruchov OY, Nieznanski K (Dec 1997). "Kinase-related protein (telokin) is phosphorylated by smooth-muscle myosin light-chain kinase and modulates the kinase activity". The Biochemical Journal. 328. 328 (2): 425–30. doi:10.1042/bj3280425. PMC   1218937 . PMID   9371697.
  3. ↑ Nieznanski K, Sobieszek A (Feb 1997). "Telokin (kinase-related protein) modulates the oligomeric state of smooth-muscle myosin light-chain kinase and its interaction with myosin filaments". The Biochemical Journal. 322. 322 (1): 65–71. doi:10.1042/bj3220065. PMC   1218159 . PMID   9078244.
  4. ↑ Ito M, Dabrowska R, Guerriero V, Hartshorne DJ (Aug 1989). "Identification in turkey gizzard of an acidic protein related to the C-terminal portion of smooth muscle myosin light chain kinase". The Journal of Biological Chemistry. 264 (24): 13971–4. PMID   2760053.
  5. ↑ "Reaction information for EC 2.7.11.18 - myosin-light-chain kinase". BRENDA.
  6. ↑ "KM values for EC 2.7.11.18 - myosin-light-chain kinase". BRENDA.
  7. 1 2 Herring BP, Lyons GE, Hoggatt AM, Gallagher PJ (Jan 2001). "Telokin expression is restricted to smooth muscle tissues during mouse development". American Journal of Physiology. Cell Physiology. 280 (1): C12-21. doi:10.1152/ajpcell.2001.280.1.c12. PMC   2860107 . PMID   11121372.
  8. 1 2 3 4 Shirinsky VP, Vorotnikov AV, Birukov KG, Nanaev AK, Collinge M, Lukas TJ, Sellers JR, Watterson DM (1993). "A kinase-related protein stabilizes unphosphorylated smooth muscle myosin minifilaments in the presence of ATP". J. Biol. Chem. 268 (22): 16578–83. PMID   8344938.
  9. 1 2 3 Gallagher PJ, Herring BP (Dec 1991). "The carboxyl terminus of the smooth muscle myosin light chain kinase is expressed as an independent protein, telokin". The Journal of Biological Chemistry. 266 (35): 23945–52. PMC   2836763 . PMID   1748667.
  10. ↑ Collinge M, Matrisian PE, Zimmer WE, Shattuck RL, Lukas TJ, Van Eldik LJ, Watterson DM (May 1992). "Structure and expression of a calcium-binding protein gene contained within a calmodulin-regulated protein kinase gene". Molecular and Cellular Biology. 12 (5): 2359–71. doi:10.1128/mcb.12.5.2359. PMC   364408 . PMID   1373815.
  11. 1 2 Vorotnikov AV (1997). "Kinase-related protein: a smooth muscle myosin-binding protein". Int. J. Biochem. Cell Biol. 29 (5): 727–30. doi:10.1016/S1357-2725(96)00156-2. PMID   9251240.
  12. ↑ Herring BP, El-Mounayri O, Gallagher PJ, Yin F, Zhou J (Nov 2006). "Regulation of myosin light chain kinase and telokin expression in smooth muscle tissues". American Journal of Physiology. Cell Physiology. 291 (5): C817-27. doi:10.1152/ajpcell.00198.2006. PMC   2836780 . PMID   16774989.
  13. ↑ Léguillette R, Laviolette M, Bergeron C, Zitouni N, Kogut P, Solway J, Kachmar L, Hamid Q, Lauzon AM (Feb 2009). "Myosin, transgelin, and myosin light chain kinase: expression and function in asthma". American Journal of Respiratory and Critical Care Medicine. 179 (3): 194–204. doi:10.1164/rccm.200609-1367OC. PMC   2633053 . PMID   19011151.
  14. ↑ Graham WV, Wang F, Clayburgh DR, Cheng JX, Yoon B, Wang Y, Lin A, Turner JR (Sep 2006). "Tumor necrosis factor-induced long myosin light chain kinase transcription is regulated by differentiation-dependent signaling events. Characterization of the human long myosin light chain kinase promoter". The Journal of Biological Chemistry. 281 (36): 26205–15. doi: 10.1074/jbc.M602164200 . PMID   16835238.
  15. ↑ Léveillé N, Fournier A, Labrie C (Apr 2009). "Androgens down-regulate myosin light chain kinase in human prostate cancer cells". The Journal of Steroid Biochemistry and Molecular Biology. 114 (3–5): 174–9. doi:10.1016/j.jsbmb.2009.02.002. PMID   19429448. S2CID   1695778.
  16. ↑ Wang L, Guo DC, Cao J, Gong L, Kamm KE, Regalado E, Li L, Shete S, He WQ, Zhu MS, Offermanns S, Gilchrist D, Elefteriades J, Stull JT, Milewicz DM (Nov 2010). "Mutations in myosin light chain kinase cause familial aortic dissections". American Journal of Human Genetics. 87 (5): 701–7. doi:10.1016/j.ajhg.2010.10.006. PMC   2978973 . PMID   21055718.
  17. ↑ "variant VAR_065577". UniProtKB/SwissProt.
  18. ↑ Madden JA, Dantuma MW, Sorokina EA, Weihrauch D, Kleinman JG (Jun 2008). "Telokin expression and the effect of hypoxia on its phosphorylation status in smooth muscle cells from small and large pulmonary arteries". American Journal of Physiology. Lung Cellular and Molecular Physiology. 294 (6): L1166–73. doi:10.1152/ajplung.00375.2007. PMID   18375742.
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