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A splicing alteration of 4.1R pre-mRNA generates 2 protein isoforms with distinct assembly to spindle poles in

François Delhommeau1, Corinne Vasseur-Godbillon, Philippe Leclerc

  • 1Institut National de la Santé et de la Recherche Médicale, U 473 and U 488, Le Kremlin-Bicêtre, France.

Blood
|September 20, 2002
PubMed

Insights

The C-terminal region of protein 4.1R is crucial for its function in cell division. A specific isoform, CO.2, maintains this function by binding to spindle poles, unlike CO.1.

Area of Science:

  • Cell Biology
  • Molecular Biology
  • Biochemistry

Background:

  • The C-terminal region of erythroid cytoskeletal protein 4.1R (encoded by exons 20 and 21) binds to nuclear mitotic apparatus protein (NuMA), essential for mitotic spindle formation and stability.
  • Splicing mutations in 4.1R produce two isoforms: CO.1 (lacking most of exon 20 peptide) and CO.2 (lacking exon 20 C-terminal sequence but retaining exon 21 sequence).
  • Both isoforms are expressed in red blood cells and associate with the membrane skeleton.

Purpose of the Study:

  • To investigate whether the 4.1R isoforms, CO.1 and CO.2, can fulfill the mitotic functions of 4.1R in dividing cells.
  • To determine the role of the C-terminal region of 4.1R in its association with the mitotic spindle and centrosomes.
  • To explore the potential functional redundancy between 4.1R and its homolog 4.1G in mitotic cells.

Main Methods:

  • Analysis of protein localization in erythroid and lymphoid mutated cells.
  • In vitro interaction studies between 4.1R isoforms and NuMA.
  • Observation of protein localization under microtubule-destabilizing conditions.
  • Investigation of 4.1G expression and its C-terminal interaction with NuMA.

Main Results:

  • CO.2, but not CO.1, assembles to spindle poles and colocalizes with NuMA in mutated cells.
  • Neither CO.1 nor CO.2 isoforms interact with NuMA in vitro.
  • CO.2 consistently localizes to centrosomes, even under microtubule-destabilizing conditions.
  • The ubiquitous homolog 4.1G is present in cells and its C-terminal region binds NuMA effectively.

Conclusions:

  • The stability of 4.1R's association with centrosomes requires an intact C-terminal end, potentially for proper protein conformation or binding to an unknown centrosome-cytoskeletal network.
  • Mitotic spindles likely contain a mixture of 4.1R and 4.1G proteins.
  • Coexpression of 4.1R and 4.1G at spindle poles may indicate functional redundancy in mitotic cells.

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