Abstract
BackgroundThe biogenesis of spliceosomal snRNPs takes place in both the cytoplasm where Sm core proteins are added and snRNAs are modified at the 5′ and 3′ termini and in the nucleus where snRNP-specific proteins associate. U1 snRNP consists of U1 snRNA, seven Sm proteins and three snRNP-specific proteins, U1-70K, U1A, and U1C. It has been shown previously that after import to the nucleus U2 and U4/U6 snRNP-specific proteins first appear in Cajal bodies (CB) and then in splicing speckles. In addition, in cells grown under normal conditions U2, U4, U5, and U6 snRNAs/snRNPs are abundant in CBs. Therefore, it has been proposed that the final assembly of these spliceosomal snRNPs takes place in this nuclear compartment. In contrast, U1 snRNA in both animal and plant cells has rarely been found in this nuclear compartment.Methodology/Principal FindingsHere, we analysed the subnuclear distribution of Arabidopsis U1 snRNP-specific proteins fused to GFP or mRFP in transiently transformed Arabidopsis protoplasts. Irrespective of the tag used, U1-70K was exclusively found in the nucleus, whereas U1A and U1C were equally distributed between the nucleus and the cytoplasm. In the nucleus all three proteins localised to CBs and nucleoli although to different extent. Interestingly, we also found that the appearance of the three proteins in nuclear speckles differ significantly. U1-70K was mostly found in speckles whereas U1A and U1C in ∼90% of cells showed diffuse nucleoplasmic in combination with CBs and nucleolar localisation.Conclusions/SignificanceOur data indicate that CBs and nucleolus are involved in the maturation of U1 snRNP. Differences in nuclear accumulation and distribution between U1-70K and U1A and U1C proteins may indicate that either U1-70K or U1A and U1C associate with, or is/are involved, in other nuclear processes apart from pre-mRNA splicing.
Highlights
Pre-mRNA splicing is mediated by the spliceosome, a dynamic macromolecular complex which assembles anew on each intron
We could previously show that transient expression of U2 small nuclear ribonucleoprotein particles (snRNPs)-specific proteins in Arabidopsis protoplasts results in their correct localisation in the nucleus
It has been shown that U2B0 and U2A9 proteins localise in a diffuse/speckled nucleoplasmic pattern, with the majority of cells showing localisation in Cajal bodies (CB) and to a lesser extent in nucleoli [34]
Summary
Pre-mRNA splicing is mediated by the spliceosome, a dynamic macromolecular complex which assembles anew on each intron. Each snRNP is composed of one uridine-rich snRNAs and the seven common Sm (or Lsm, in the case of U6 snRNP) proteins, B/B9, D1, D2, D3, E, F and G. In addition to the Sm/Lsm proteins, each snRNP contains particle-specific proteins (see below). The biogenesis of spliceosomal snRNPs takes place in both the cytoplasm where Sm core proteins are added and snRNAs are modified at the 59 and 39 termini and in the nucleus where snRNP-specific proteins associate. It has been shown previously that after import to the nucleus U2 and U4/U6 snRNP-specific proteins first appear in Cajal bodies (CB) and in splicing speckles. In cells grown under normal conditions U2, U4, U5, and U6 snRNAs/snRNPs are abundant in CBs. it has been proposed that the final assembly of these spliceosomal snRNPs takes place in this nuclear compartment. U1 snRNA in both animal and plant cells has rarely been found in this nuclear compartment
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