Abstract
Gametogenesis and embryogenesis are dynamic developmental stages marked by extensive modifications in the organization of the genome and nuclear architecture. In the literature it is conveyed that only B-type lamins are required in these early stages of development and that A-type lamins are not present or required until differentiation of specific cell types associated with specialized tissue is initiated. To assess the presence of nuclear structures that are putatively involved in genome regulation, we investigated the distribution of lamin proteins throughout the early stages of porcine embryonic development, using testes tissue sections, oocytes and in-vitro fertilized (IVF) porcine embryos and employing anti-lamin antibodies. We have shown that anti-lamin A staining is present at the one-cell, two-cell, four-cell, and six- to eight-cell stages of early porcine embryo development, but diminishes at the morulae and blastocyst stages. Large intranuclear anti-lamin A foci are prominent in the early preimplantation stages. Both anti-lamin A/C and anti-lamin B staining were clearly present in all embryonic stages. Immature porcine oocytes revealed lamin rings using the monoclonal anti-lamin A/C antibody and many immature oocytes exhibited a pale rim staining pattern with anti-lamin A antibody. A-type lamins were not observed in sperm precursor cells. Thus, we have shown that A-type lamins and B-type lamins are present at the nuclear envelope in very early porcine embryos and that lamin A is also found in large intranuclear aggregates in two-cell to eight-cell embryos but is lacking from later embryonic stages.
Similar content being viewed by others
References
Abeydeera LR, Day BN (1997) Fertilization and subsequent development in vitro of pig oocytes inseminated in a modified Tris-buffered medium with frozen–thawed ejaculated spermatozoa. Biol Reprod 57: 729-34.
Abeydeera LR, Wang W-H, Cantley TC et al. (2000) Development and viability of pig oocytes matured in a protein-free medium containing epidermal growth factor. Theriogenology 54: 787-97.
Aebi U, Cohn J, Buhle L, Gerace L (1986) The nuclear lamina is a meshwork of intermediate-type filaments. Nature 323: 560-64.
Alsheimer M, Benavente R (1996) Change of karyoskeleton during mammalian spermatogenesis: expression pattern of nuclear lamin C2 and its regulation. Exp Cell Res 228: 181-88.
Alsheimer M, Fecher E, Benavente R (1998) Nuclear envelope remodelling during rat spermogenesis: distribution and expression pattern of LAP2/thymopoietins. J Cell Sci 111: 2227-234.
Alsheimer M, von Glasenapp E, Hock R, Benavente R (1999) Architecture of the nuclear periphery of rat pachytene spermatocytes: distribution of nuclear envelope proteins in relation to synaptonemal complex attachment sites. Mol Biol Cell 10: 1235-245.
Alsheimer M, Liebe B, Sewell L, Stewart CL, Scherthan H, Benavente R (2004) Disruption of spermatogenesis in mice lacking A-type lamins. J Cell Sci 117: 1173-178.
Barboro P, D’Arrigo C, Diaspro A et al. (2002) Unraveling the organization of the internal nuclear matrix: RNA-dependent anchoring of NuMA to a lamin scaffold. Exp Cell Res 279: 202-18.
Bridger JM, Kill IR, O’Farrell M, Hutchison CJ (1993) Internal lamin structures within G1 nuclei of human dermal fibroblasts. J Cell Sci 104: 297-06.
Bridger JM, Foeger N, Kill IR, Herrmann H (2006) The nuclear lamina — a structural framework involved in genome organization. FEBS J (In press).
Christians E, Boiani M, Garagna S et al. (1999) Gene expression and chromatin organization during mouse oocyte growth. Dev Biol 207: 76-5.
Crozet N, Motlik J, Szollosi D (1981) Nucleolar fine structure and RNA synthesis in porcine oocytes during early stages of antrum formation. Biol Cell 41: 35-2.
Degrouard J, Hozak P, Heyman Y, Flechon JE (2004) Nucleoskeleton of early bovine embryos and differentiated somatic cells: an ultrastructural and immunocytochemical comparison. Histochem Cell Biol 121: 441-51.
Fisher DZ, Chaudhary N, Blobel G (1986) cDNA sequencing of nuclear lamins A and C reveals primary and secondary structural homology to intermediate filament proteins. Proc Natl Acad Sci USA 83: 6450-454.
Foster HA, Bridger JM (2005) The genome and the nucleus: a marriage made by evolution. Genome organization and nuclear architecture. Chromosoma 114: 212-19.
Foster HA, Abeydeera LR, Griffin DK, Bridger JM (2005) Non-random chromosome positioning in mammalian sperm nuclei, with migration of the sex chromosomes during late spermatogenesis. J Cell Sci 118: 1811-820.
Freitag M, Dopke HH, Niemann H, Elsaesser F (1991) 3H-uridine incorporation in early porcine embryos. Mol Reprod Dev 29: 124-28.
Furukawa K, Hotta Y (1993) cDNA cloning of a germ cell specific lamin B3 from mouse spermatocytes and analysis of its function by ectopic expression in somatic cells. EMBO J 12: 97-06.
Furukawa K, Inagaki H, Hotta Y (1994) Identification and cloning of an mRNA coding for a germ cell-specific A-type lamin in mice. Exp Cell Res 212: 426-30.
Goldman RD, Gruenbaum Y, Moir RD, Shumaker DK, Spann TP (2002) Nuclear lamins: building blocks of nuclear architecture. Genes Dev 16: 533-47.
Hall VJ, Cooney MA, Shanahan P, Tecirlioglu RT, Ruddock NT, French AJ (2005) Nuclear lamin antigen and messenger RNA expression in bovine in vitro produced and nuclear transfer embryos. Mol Reprod Dev 72: 471-82.
Hennekes H, Nigg EA (1994) The role of isoprenylation in membrane attachment of nuclear lamins. A single point mutation prevents proteolytic cleavage of the lamin A precursor and confers membrane binding properties. J Cell Sci 107: 1019-029.
Holtz D, Tanaka RA, Hartwig J, McKeon F (1989) The CaaX motif of lamin A functions in conjunction with the nuclear localization signal to target assembly to the nuclear envelope. Cell 59: 969-77.
Hozak P, Sasseville AM, Raymond Y, Cook PR (1995) Lamin proteins form an internal nucleoskeleton as well as a peripheral lamina in human cells. J Cell Sci 108: 635-44.
Hutchison CJ, Worman HJ (2004) A-type lamins: guardians of the soma? Nat Cell Biol 6: 1062-067.
Hutchison CJ, Bridger JM, Cox LS, Kill IR (1994) Weaving a pattern from disparate threads: lamin function in nuclear assembly and DNA replication. J Cell Sci 107: 3259-269.
Hyttel P, Laurincik J, Rosenkranz C, Rath D, Niemann H, Ochs RL, Schellander K (2000) Nucleolar proteins and ultrastructure in preimplantation porcine embryos developed in vivo. Biol Reprod 63: 1848-856.
Jarrell VL, Day BN, Prather RS (1991) The transition from maternal to zygotic control of development occurs during the 4-cell stage in the domestic pig, Sus scrofa: quantitative and qualitative aspects of protein synthesis. Biol Reprod 44: 62–68.
Jenkins H, Whitfield WG, Goldberg MW, Allen TD, Hutchison CJ (1995) Evidence for the direct involvement of lamins in the assembly of a replication competent nucleus. Acta Biochim Pol 42: 133-43.
Jeong BS, Yang X (2001) Cysteine, glutathione, and Percoll treatments improve porcine oocyte maturation and fertilization in vitro. Mol Reprod Dev 59: 330-35.
Krawetz SA (2005) Paternal contribution: new insights and future challenges. Nat Rev Genet 6: 633-42.
Lazebnik YA, Takahashi A, Moir RD et al. (1995) Studies of the lamin proteinase reveal multiple parallel biochemical pathways during apoptotic execution. Proc Natl Acad Sci USA 92: 9042-046.
Lehner CF, Stick R, Eppenberger HM, Nigg EA (1987) Differential expression of nuclear lamin proteins during chicken development. J Cell Biol 105: 577-87.
Liu J, Rolef-Ben-Shahar TR, Riemer D et al. (2000) Essential roles for Caenorhabditis elegans lamin gene in nuclear organization, cell cycle progression, and spatial organization of nuclear pore complexes. Mol Biol Cell 11: 3937-947.
Longo F, Garagna S, Merico V et al. (2003) Nuclear localization of NORs and centromeres in mouse oocytes during folliculogenesis. Mol Reprod Dev 66: 279-90.
Lourim D, Krohne G (1993) Membrane-associated lamins in Xenopus egg extracts: identification of two vesicle populations. J Cell Biol 123: 501-12.
Lourim D, Kempf A, Krohne G (1996) Characterization and quantitation of three B-type lamins in Xenopus oocytes and eggs: increase of lamin LI protein synthesis during meiotic maturation. J Cell Sci 109: 1775-785.
Mancini MA, Shan B, Nickerson JA, Penman S, Lee WH (1994) The retinoblastoma gene product is a cell cycle-dependent, nuclear matrix-associated protein. Proc Natl Acad Sci USA 91: 418-22.
Maul GG, Schatten G, Jimenez SA, Carrera AE (1987) Detection of nuclear lamin B epitopes in oocyte nuclei from mice, sea urchins, and clams using a human autoimmune serum. Dev Biol 121: 368-75.
Meier J, Campbell KH, Ford CC, Stick R, Hutchison CJ (1991) The role of lamin LIII in nuclear assembly and DNA replication, in cell-free extracts of Xenopus eggs. J Cell Sci 98: 271-79.
Moir RD, Spann TP, Herrmann H, Goldman RD (2000) Disruption of nuclear lamin organization blocks the elongation phase of DNA replication. J Cell Biol 149: 1179-192.
Nagai T, Funahashi H, Yoshioka K, Kikuchi K (2006) Update of in vitro production of porcine embryos. Front Biosci 11: 2565-573.
Newport JW, Wilson KL, Dunphy WG (1990) A lamin-independent pathway for nuclear envelope assembly. J Cell Biol 111: 2247-259.
Petters RM, Johnson BH, Reed ML, Archibong AE (1990) Glucose, glutamine and inorganic phosphate in early development of the pig embryo in vitro. J Reprod Fertil 89: 269-75.
Prather RS, Sims MM, Maul GG, First NL, Schatten G (1989) Nuclear lamin antigens are developmentally regulated during porcine and bovine embryogenesis. Biol Reprod 41: 123-32.
Pugh GE, Coates PJ, Lane EB, Raymond Y, Quinlan RA (1997) Distinct nuclear assembly pathways for lamins A and C lead to their increase during quiescence in Swiss 3T3 cells. J Cell Sci 110: 2483-493.
Rober RA, Weber K, Osborn M (1989) Differential timing of nuclear lamin A/C expression in the various organs of the mouse embryo and the young animal: a developmental study. Development 105: 365-78.
Schatten G, Maul GG, Schatten H et al. (1985) Nuclear lamins and peripheral nuclear antigens during fertilization and embryogenesis in mice and sea urchins. Proc Natl Acad Sci USA 82: 4727-731.
Schoenbeck RA, Peters MS, Rickords LF, Stumpf TT, Prather RS (1992) Characterization of deoxyribonucleic acid synthesis and the transition from maternal to embryonic control in the 4-cell porcine embryo. Biol Reprod 47: 1118-125.
Smith DE, Fisher PA (1989) Interconversion of Drosophila nuclear lamin isoforms during oogenesis, early embryogenesis, and upon entry of cultured cells into mitosis. J Cell Biol 108: 255-65.
Spann TP, Moir RD, Goldman AE, Stick R, Goldman RD (1997) Disruption of nuclear lamin organization alters the distribution of replication factors and inhibits DNA synthesis. J Cell Biol 136: 1201-212.
Spann TP, Goldman AE, Wang C, Huang S, Goldman RD (2002) Alteration of nuclear lamin organization inhibits RNA polymerase II-dependent transcription. J Cell Biol 156: 603-08.
Stokes PJ, Abeydeera LR, Leese HJ (2005) Development of porcine embryos in vivo and in vitro; evidence for embryo ‘cross talk-in vitro. Dev Biol 284: 62-1.
Sturmey RG, Leese HJ (2003) Energy metabolism in pig oocytes and early embryos. Reproduction 126: 197-04.
Taniura H, Glass C, Gerace L (1995) A chromatin binding site in the tail domain of nuclear lamins that interacts with core histones. J Cell Biol 131: 33-4.
Tomanek M, Kopecny V, Kanka J (1989) Genome reactivation in developing early pig embryos: an ultrastructural and autoradiographic analysis. Anat Embryol (Berl) 180: 309-16.
Vergnes L, Peterfy M, Bergo MO, Young SG, Reue K (2004) Lamin B1 is required for mouse development and nuclear integrity. Proc Natl Acad Sci USA 101: 10428-0433.
Viuff D, Greve T, Holm P, Callesen H, Hyttel P, Thomsen PD (2002) Activation of the ribosomal RNA genes late in the third cell cycle of porcine embryos. Biol Reprod 66: 629-34.
Zuccotti M, Piccinelli A, Giorgi Rossi P, Garagna S, Redi CA (1995) Chromatin organization during mouse oocyte growth. Mol Reprod Dev 41: 479-85.
Zuccotti M, Giorgi Rossi P, Martinez A, Garagna S, Forabosco A, Redi CA (1998) Meiotic and developmental competence of mouse antral oocytes. Biol Reprod 58: 700-04.
Author information
Authors and Affiliations
Corresponding author
Electronic Supplementary Material
Below is the link to the electronic supplementary material.
Supplementary Figure 1
Indirect immunofluorescence on porcine testes tissue. Monoclonal anti-lamin A staining 133A2 (A) polyclonal anti-lamin A C-20 (B) monoclonal anti-lamin A/C 636 (C) and polyclonal anti-lamin B C-20 (D). Heterogeneous positive anti-lamin A/C 636 staining was determined in certain nuclei (C- C-). Nuclei were counterstained with propidium iodide. The solid arrow depicts a spermatocyte nucleus, whereas the hatched arrow portrays a spermatid nucleus. Bar=10 μm. Porcine testes were incubated with sterile 40% sucrose for 3 days in order to replace the water and to allow easier sectioning when frozen. The testes tissue was cut into small pieces, frozen in hexane bath, and stored at -80°C. Sections of frozen tissue (60 μm ) were cut using a cryomicrotome (Bright 5030 microtome), and adhered to slides coated with 3% 3-aminopropyltriethoxysilane (APES). The tissue sections were stored at -80°C until required, they were then air-dried for at least 1 h prior to fixation for 10 min with ice-cold acetone. They were then rinsed three times with PBS and incubated with 10% (NCS) for 10 min, before application of the primary antibody. The tissue sections were incubated in primary antibodies mouse anti-lamin A 133A2 antibody diluted 1:400, goat anti-lamin A C-20 antibody diluted 1:100, mouse anti-lamin A/C 636 antibody and goat anti-lamin B C-20 antibody diluted 1:100, all diluted with 1% (v/v) NCS in PBS and incubated with the slide for 24 h at 4°C. The tissue sections were washed three times with PBS and incubated with the appropriate secondary antibody (donkey anti-mouse FITC diluted 1:60; Jackson Laboratory or donkey anti-goat FITC diluted 1:60; Jackson Laboratory) for 30 min at 37°C. The sections were rinsed three times in PBS prior to mounting in Vectashield anti-fade mountant (Vectorlabs) containing propidium iodide (PI) as a counterstain. (JPG 755 993)
Rights and permissions
About this article
Cite this article
Foster, H.A., Stokes, P., Forsey, K. et al. Lamins A and C are present in the nuclei of early porcine embryos, with lamin A being distributed in large intranuclear foci. Chromosome Res 15, 163–174 (2007). https://doi.org/10.1007/s10577-006-1088-8
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10577-006-1088-8