Date of Award

Fall 2014

Degree Type


Degree Name

Doctor of Philosophy (PhD)


Animal Science

First Advisor

Ryan Cabot

Committee Chair

Ryan Cabot

Committee Member 1

Zoltan Machaty

Committee Member 2

Kolapo Ajuwon

Committee Member 3

Shihuan Kuang

Committee Member 4

Ann Kirchmaier


Intracellular communication between the nucleus and cytoplasm is critically important for coordinating cellular events during embryogenesis. The karyopherin α/β heterodimer is an intracellular nuclear trafficking system that mediates nuclear import of proteins that bear classical nuclear localization signals (NLSs). Seven karyopherin α subtypes have been identified in the domestic pig, and while each of these karyopherin α subtypes is able to bind to a nuclear localization signal, individual karyopherin α subtypes have been shown to transport specific NLS-bearing proteins. The objective of this study was to determine the developmental requirements of karyopherin α subtypes (KPNAs) during cleavage development in porcine embryos. The purpose of this dissertation was to test the hypothesis that the karyopherin α/β heterodimer-mediated nuclear trafficking pathway serves regulatory roles during cleavage development by selectively partitioning intracellular cargoes, thereby affecting epigenetic modifications, transcription, and embryo developmental potential. We tested our hypothesis via a combination of a series of in vivo and in vitro assays. Our microinjection assay revealed that POU domain, class 3, transcription factor 2 (BRN2, also referred to as POU3F2) adopts a nuclear localization in all nuclei through the 4-cell stage of development, while only a subset of blastomeres in 8-cell stage embryos possess nuclear BRN2. Octamer-binding transcription factor 4 (OCT4) adopts a nuclear localization in all nuclei prior to the 2-cell stage of development, whereas OCT4 is undetectable in nuclei at the 4-cell stage. In vitro binding assays showed that both BRN2 and OCT4 are able to bind with multiple porcine karyopherin α subtypes. Moreover, we tested the impact of KPNA1-depletion in the intracellular localization of BRN2 and the embryo developmental competence via a series of co-microinjection assays. Our results showed that GFP-BRN2 accumulation was significantly reduced in the nuclei of KPNA1-depleted embryos, and KPNA1-depleted embryos possessed significantly fewer nuclei as well as a reduced proportion with the capacity to develop to the 8-cell stage and beyond as compared with the control embryos.^ In summary, the data discussed in this dissertation provide more evidence to support the conclusion that discrete classes of NLS-bearing nuclear proteins may be preferentially imported by individual karyopherin α subtypes. Our data also indicate that KPNA1 might be involved in the transport of transcription factor BRN2 in 4-cell stage embryos. Furthermore, the data suggest that KPNA1 may serve a critical role of partitioning intracellular cargoes that are directly involved in zygotic genome activation, or that direct development immediately after zygotic genome activation, hence affecting early cleavage embryo development.

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Agriculture Commons