Knockdown of Splicing Complex Protein PCBP2 Reduces Extravillous Trophoblast Differentiation Through Transcript...

Area of Science:

• Reproductive Biology
• Molecular Genetics
• RNA Biology

Background:

• Mutations in LINC-HELLP (HELLPAR) non-coding RNA are linked to familial HELLP syndrome.
• These mutations impair extravillous trophoblast (EVT) differentiation and RNA splicing protein binding.
• The roles of PCBP1, PCBP2, and YBX1 in EVT invasion and placentation require further investigation.

Purpose of the Study:

• To investigate the involvement of PCBP1, PCBP2, and YBX1 in regulating EVT invasion during placentation.
• To determine if alternative mRNA splicing mediated by these proteins influences EVT differentiation.
• To elucidate the mechanism by which PCBP2 affects EVT function and its potential link to HELLP syndrome.

Main Methods:

• Utilized in vitro (HTR-8/SVneo cells) and ex vivo (human placental explants) models.
• Employed siRNA-mediated gene silencing to downregulate PCBP1, PCBP2, and YBX1.
• Performed Transwell invasion assays, proliferation assays, and whole transcriptome sequencing (RNA-seq).

Main Results:

• Knockdown of PCBP2, and to a lesser extent PCBP1 and YBX1, inhibited EVT differentiation and invasion.
• PCBP2 knockdown in placental explants reduced EVT outgrowth by approximately 80% and increased proliferation in remaining EVTs.
• RNA-seq revealed PCBP2 silencing primarily altered alternative splicing, not mRNA expression, affecting genes involved in cellular organization, function, and cell cycle.

Conclusions:

• PCBP2 is a critical regulator of EVT differentiation and invasion, essential for proper placental development.
• Dysregulated binding of PCBP2 to LINC-HELLP may contribute to placental dysfunction observed in HELLP syndrome.
• Alternative splicing orchestrated by PCBP2 plays a key role in EVT functional specialization for decidual invasion.