Skip to main content
NCBI home page
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1985 Nov;5(11):3058–3068. doi: 10.1128/mcb.5.11.3058

Characterization of the myosin light-chain-2 gene of Drosophila melanogaster.

V P Parker, S Falkenthal, N Davidson
PMCID: PMC369119  PMID: 3018498

Abstract

Recombinant DNA clones encoding the Drosophila melanogaster homolog of the vertebrate myosin light-chain-2 (MLC-2) gene have been isolated. This single-copy gene maps to the chromosomal locus 99E. The nucleotide sequence was determined for a 3.4-kilobase genomic fragment containing the gene and for two MLC-2 cDNA clones generated from late pupal mRNA. Comparison of these sequences shows that the gene contains two introns, the positions of which are conserved in the corresponding rat sequence. Extension of a primer homologous to the mRNA reveals two start sites for transcription 12 nucleotides apart. The sequence TATA is not present ahead of the mRNA cap site. There are two major sites of poly(A) addition separated by 356 nucleotides. The protein sequence derived from translation of the cDNA sequence shows a high degree of homology with that for the DTNB myosin light chain (MLC-2) of chicken. A lower degree of sequence homology was seen in comparisons with other evolutionarily related calcium-binding proteins. RNA blots show high levels of expression of several transcripts during the developmental time stages when muscle is being produced. In vitro translation of hybrid-selected RNA produces two polypeptides which comigrate on two-dimensional gels with proteins from Drosophila actomyosin, although the cDNA sequence reveals only one 26-kilodalton primary translation product.

Full text

PDF
3058

Images in this article

3061Image
on p.3061
3062Image
on p.3062
3064Image
on p.3064
3065Image
on p.3065

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Basi G. S., Boardman M., Storti R. V. Alternative splicing of a Drosophila tropomyosin gene generates muscle tropomyosin isoforms with different carboxy-terminal ends. Mol Cell Biol. 1984 Dec;4(12):2828–2836. doi: 10.1128/mcb.4.12.2828. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Benton W. D., Davis R. W. Screening lambdagt recombinant clones by hybridization to single plaques in situ. Science. 1977 Apr 8;196(4286):180–182. doi: 10.1126/science.322279. [DOI] [PubMed] [Google Scholar]
  3. Bergsma D. J., Chang K. S., Schwartz R. J. Novel chicken actin gene: third cytoplasmic isoform. Mol Cell Biol. 1985 May;5(5):1151–1162. doi: 10.1128/mcb.5.5.1151. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Berk A. J., Sharp P. A. Sizing and mapping of early adenovirus mRNAs by gel electrophoresis of S1 endonuclease-digested hybrids. Cell. 1977 Nov;12(3):721–732. doi: 10.1016/0092-8674(77)90272-0. [DOI] [PubMed] [Google Scholar]
  5. Bolivar F., Rodriguez R. L., Greene P. J., Betlach M. C., Heyneker H. L., Boyer H. W., Crosa J. H., Falkow S. Construction and characterization of new cloning vehicles. II. A multipurpose cloning system. Gene. 1977;2(2):95–113. [PubMed] [Google Scholar]
  6. Breathnach R., Chambon P. Organization and expression of eucaryotic split genes coding for proteins. Annu Rev Biochem. 1981;50:349–383. doi: 10.1146/annurev.bi.50.070181.002025. [DOI] [PubMed] [Google Scholar]
  7. Brosius J., Dull T. J., Noller H. F. Complete nucleotide sequence of a 23S ribosomal RNA gene from Escherichia coli. Proc Natl Acad Sci U S A. 1980 Jan;77(1):201–204. doi: 10.1073/pnas.77.1.201. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Brosius J., Palmer M. L., Kennedy P. J., Noller H. F. Complete nucleotide sequence of a 16S ribosomal RNA gene from Escherichia coli. Proc Natl Acad Sci U S A. 1978 Oct;75(10):4801–4805. doi: 10.1073/pnas.75.10.4801. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Collins J. H. Homology of myosin DTNB light chain with alkali light chains, troponin C and parvalbumin. Nature. 1976 Feb 26;259(5545):699–700. doi: 10.1038/259699a0. [DOI] [PubMed] [Google Scholar]
  10. Dierks P., van Ooyen A., Cochran M. D., Dobkin C., Reiser J., Weissmann C. Three regions upstream from the cap site are required for efficient and accurate transcription of the rabbit beta-globin gene in mouse 3T6 cells. Cell. 1983 Mar;32(3):695–706. doi: 10.1016/0092-8674(83)90055-7. [DOI] [PubMed] [Google Scholar]
  11. Falkenthal S., Parker V. P., Davidson N. Developmental variations in the splicing pattern of transcripts from the Drosophila gene encoding myosin alkali light chain result in different carboxyl-terminal amino acid sequences. Proc Natl Acad Sci U S A. 1985 Jan;82(2):449–453. doi: 10.1073/pnas.82.2.449. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Falkenthal S., Parker V. P., Mattox W. W., Davidson N. Drosophila melanogaster has only one myosin alkali light-chain gene which encodes a protein with considerable amino acid sequence homology to chicken myosin alkali light chains. Mol Cell Biol. 1984 May;4(5):956–965. doi: 10.1128/mcb.4.5.956. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Favaloro J., Treisman R., Kamen R. Transcription maps of polyoma virus-specific RNA: analysis by two-dimensional nuclease S1 gel mapping. Methods Enzymol. 1980;65(1):718–749. doi: 10.1016/s0076-6879(80)65070-8. [DOI] [PubMed] [Google Scholar]
  14. Fyrberg E. A., Bond B. J., Hershey N. D., Mixter K. S., Davidson N. The actin genes of Drosophila: protein coding regions are highly conserved but intron positions are not. Cell. 1981 Apr;24(1):107–116. doi: 10.1016/0092-8674(81)90506-7. [DOI] [PubMed] [Google Scholar]
  15. Fyrberg E. A., Kindle K. L., Davidson N., Kindle K. L. The actin genes of Drosophila: a dispersed multigene family. Cell. 1980 Feb;19(2):365–378. doi: 10.1016/0092-8674(80)90511-5. [DOI] [PubMed] [Google Scholar]
  16. Garfinkel M. D., Pruitt R. E., Meyerowitz E. M. DNA sequences, gene regulation and modular protein evolution in the Drosophila 68C glue gene cluster. J Mol Biol. 1983 Aug 25;168(4):765–789. doi: 10.1016/s0022-2836(83)80074-6. [DOI] [PubMed] [Google Scholar]
  17. Ghosh P. K., Reddy V. B., Piatak M., Lebowitz P., Weissman S. M. Determination of RNA sequences by primer directed synthesis and sequencing of their cDNA transcripts. Methods Enzymol. 1980;65(1):580–595. doi: 10.1016/s0076-6879(80)65061-7. [DOI] [PubMed] [Google Scholar]
  18. Goodman M., Pechère J. F., Haiech J., Demaille J. G. Evolutionary diversification of structure and function in the family of intracellular calcium-binding proteins. J Mol Evol. 1979 Nov;13(4):331–352. doi: 10.1007/BF01731373. [DOI] [PubMed] [Google Scholar]
  19. Grosschedl R., Birnstiel M. L. Identification of regulatory sequences in the prelude sequences of an H2A histone gene by the study of specific deletion mutants in vivo. Proc Natl Acad Sci U S A. 1980 Mar;77(3):1432–1436. doi: 10.1073/pnas.77.3.1432. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Grunstein M., Hogness D. S. Colony hybridization: a method for the isolation of cloned DNAs that contain a specific gene. Proc Natl Acad Sci U S A. 1975 Oct;72(10):3961–3965. doi: 10.1073/pnas.72.10.3961. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Hunkapiller M., Kent S., Caruthers M., Dreyer W., Firca J., Giffin C., Horvath S., Hunkapiller T., Tempst P., Hood L. A microchemical facility for the analysis and synthesis of genes and proteins. Nature. 1984 Jul 12;310(5973):105–111. doi: 10.1038/310105a0. [DOI] [PubMed] [Google Scholar]
  22. Jakes R., Northrop F., Kendrick-Jones J. Calcium binding regions of myosin 'regulatory' light chains. FEBS Lett. 1976 Nov;70(1):229–234. doi: 10.1016/0014-5793(76)80763-6. [DOI] [PubMed] [Google Scholar]
  23. Karlik C. C., Mahaffey J. W., Coutu M. D., Fyrberg E. A. Organization of contractile protein genes within the 88F subdivision of the D. melanogaster third chromosome. Cell. 1984 Jun;37(2):469–481. doi: 10.1016/0092-8674(84)90377-5. [DOI] [PubMed] [Google Scholar]
  24. Keller E. B., Noon W. A. Intron splicing: a conserved internal signal in introns of animal pre-mRNAs. Proc Natl Acad Sci U S A. 1984 Dec;81(23):7417–7420. doi: 10.1073/pnas.81.23.7417. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Kozak M. Possible role of flanking nucleotides in recognition of the AUG initiator codon by eukaryotic ribosomes. Nucleic Acids Res. 1981 Oct 24;9(20):5233–5252. doi: 10.1093/nar/9.20.5233. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Kretsinger R. H. Structure and evolution of calcium-modulated proteins. CRC Crit Rev Biochem. 1980;8(2):119–174. doi: 10.3109/10409238009105467. [DOI] [PubMed] [Google Scholar]
  27. Legerski R. J., Hodnett J. L., Gray H. B., Jr Extracellular nucleases of pseudomonas BAL 31. III. Use of the double-strand deoxyriboexonuclease activity as the basis of a convenient method for the mapping of fragments of DNA produced by cleavage with restriction enzymes. Nucleic Acids Res. 1978 May;5(5):1445–1464. doi: 10.1093/nar/5.5.1445. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Maita T., Chen J. I., Matsuda G. Amino-acid sequence of the 20 000-molecular-weight light chain of chicken gizzard-muscle myosin. Eur J Biochem. 1981 Jul;117(2):417–424. doi: 10.1111/j.1432-1033.1981.tb06354.x. [DOI] [PubMed] [Google Scholar]
  29. Matsuda G., Maita T., Kato Y., Chen J. I., Umegane T. Amino acid sequences of the cardiac L-2A, L-2B and gizzard 17 000-Mr light chains of chicken muscle myosin. FEBS Lett. 1981 Dec 7;135(2):232–236. doi: 10.1016/0014-5793(81)80789-2. [DOI] [PubMed] [Google Scholar]
  30. Maxam A. M., Gilbert W. Sequencing end-labeled DNA with base-specific chemical cleavages. Methods Enzymol. 1980;65(1):499–560. doi: 10.1016/s0076-6879(80)65059-9. [DOI] [PubMed] [Google Scholar]
  31. McLauchlan J., Gaffney D., Whitton J. L., Clements J. B. The consensus sequence YGTGTTYY located downstream from the AATAAA signal is required for efficient formation of mRNA 3' termini. Nucleic Acids Res. 1985 Feb 25;13(4):1347–1368. doi: 10.1093/nar/13.4.1347. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Messing J., Vieira J. A new pair of M13 vectors for selecting either DNA strand of double-digest restriction fragments. Gene. 1982 Oct;19(3):269–276. doi: 10.1016/0378-1119(82)90016-6. [DOI] [PubMed] [Google Scholar]
  33. Mogami K., Fujita S. C., Hotta Y. Identification of Drosophila indirect flight muscle myofibrillar proteins by means of two-dimensional electrophoresis. J Biochem. 1982 Feb;91(2):643–650. doi: 10.1093/oxfordjournals.jbchem.a133736. [DOI] [PubMed] [Google Scholar]
  34. Nabeshima Y., Fujii-Kuriyama Y., Muramatsu M., Ogata K. Alternative transcription and two modes of splicing results in two myosin light chains from one gene. Nature. 1984 Mar 22;308(5957):333–338. doi: 10.1038/308333a0. [DOI] [PubMed] [Google Scholar]
  35. Nudel U., Calvo J. M., Shani M., Levy Z. The nucleotide sequence of a rat myosin light chain 2 gene. Nucleic Acids Res. 1984 Sep 25;12(18):7175–7186. doi: 10.1093/nar/12.18.7175. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. O'Farrell P. H. High resolution two-dimensional electrophoresis of proteins. J Biol Chem. 1975 May 25;250(10):4007–4021. [PMC free article] [PubMed] [Google Scholar]
  37. Periasamy M., Strehler E. E., Garfinkel L. I., Gubits R. M., Ruiz-Opazo N., Nadal-Ginard B. Fast skeletal muscle myosin light chains 1 and 3 are produced from a single gene by a combined process of differential RNA transcription and splicing. J Biol Chem. 1984 Nov 10;259(21):13595–13604. [PubMed] [Google Scholar]
  38. Poncz M., Solowiejczyk D., Ballantine M., Schwartz E., Surrey S. "Nonrandom" DNA sequence analysis in bacteriophage M13 by the dideoxy chain-termination method. Proc Natl Acad Sci U S A. 1982 Jul;79(14):4298–4302. doi: 10.1073/pnas.79.14.4298. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Proudfoot N. J., Brownlee G. G. 3' non-coding region sequences in eukaryotic messenger RNA. Nature. 1976 Sep 16;263(5574):211–214. doi: 10.1038/263211a0. [DOI] [PubMed] [Google Scholar]
  40. Ricciardi R. P., Miller J. S., Roberts B. E. Purification and mapping of specific mRNAs by hybridization-selection and cell-free translation. Proc Natl Acad Sci U S A. 1979 Oct;76(10):4927–4931. doi: 10.1073/pnas.76.10.4927. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Roark M., Mahoney P. A., Graham M. L., Lengyel J. A. Blastoderm-differential and blastoderm-specific genes of Drosophila melanogaster. Dev Biol. 1985 Jun;109(2):476–488. doi: 10.1016/0012-1606(85)90473-7. [DOI] [PubMed] [Google Scholar]
  42. Robert B., Daubas P., Akimenko M. A., Cohen A., Garner I., Guenet J. L., Buckingham M. A single locus in the mouse encodes both myosin light chains 1 and 3, a second locus corresponds to a related pseudogene. Cell. 1984 Nov;39(1):129–140. doi: 10.1016/0092-8674(84)90198-3. [DOI] [PubMed] [Google Scholar]
  43. Rozek C. E., Davidson N. Drosophila has one myosin heavy-chain gene with three developmentally regulated transcripts. Cell. 1983 Jan;32(1):23–34. doi: 10.1016/0092-8674(83)90493-2. [DOI] [PubMed] [Google Scholar]
  44. Sanchez F., Tobin S. L., Rdest U., Zulauf E., McCarthy B. J. Two Drosophila actin genes in detail. Gene structure, protein structure and transcription during development. J Mol Biol. 1983 Feb 5;163(4):533–551. doi: 10.1016/0022-2836(83)90111-0. [DOI] [PubMed] [Google Scholar]
  45. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Snyder M., Hunkapiller M., Yuen D., Silvert D., Fristrom J., Davidson N. Cuticle protein genes of Drosophila: structure, organization and evolution of four clustered genes. Cell. 1982 Jul;29(3):1027–1040. doi: 10.1016/0092-8674(82)90466-4. [DOI] [PubMed] [Google Scholar]
  47. Suzuyama Y., Umegane T., Maita T., Matsuda G. The amino acid sequence of the L-2 light chain of chicken skeletal muscle myosin. Hoppe Seylers Z Physiol Chem. 1980;361(2):119–127. doi: 10.1515/bchm2.1980.361.1.119. [DOI] [PubMed] [Google Scholar]
  48. Ueyama H., Hamada H., Battula N., Kakunaga T. Structure of a human smooth muscle actin gene (aortic type) with a unique intron site. Mol Cell Biol. 1984 Jun;4(6):1073–1078. doi: 10.1128/mcb.4.6.1073. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Vieira J., Messing J. The pUC plasmids, an M13mp7-derived system for insertion mutagenesis and sequencing with synthetic universal primers. Gene. 1982 Oct;19(3):259–268. doi: 10.1016/0378-1119(82)90015-4. [DOI] [PubMed] [Google Scholar]
  50. Weaver R. F., Weissmann C. Mapping of RNA by a modification of the Berk-Sharp procedure: the 5' termini of 15 S beta-globin mRNA precursor and mature 10 s beta-globin mRNA have identical map coordinates. Nucleic Acids Res. 1979 Nov 10;7(5):1175–1193. doi: 10.1093/nar/7.5.1175. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Zakut R., Shani M., Givol D., Neuman S., Yaffe D., Nudel U. Nucleotide sequence of the rat skeletal muscle actin gene. Nature. 1982 Aug 26;298(5877):857–859. doi: 10.1038/298857a0. [DOI] [PubMed] [Google Scholar]

Articles from Molecular and Cellular Biology are provided here courtesy of Taylor & Francis

RESOURCES