Molecular Machines and Tissue Architecture

Our lab studies centrosomes, non-membrane bound organelles that serve as the cell’s main microtubule (MT) organizing center (MTOC). Our efforts are focused on an outstanding question in the field – how does the centrosome tune its MTOC activity to support different functions? We use a combination of genetics, molecular biology, CRISPR gene editing, super-resolution microscopy, and other cell biological approaches. Most importantly, we employ the model organism Drosophila in combination with a powerful method of generating new alleles to investigate tissue specific functions. This approach has uniquely positioned my lab to discover unappreciated roles for centrosome proteins and is guiding our current research aimed to explore the etiology of centrosome- and cilia-related human diseases such as microcephaly and infertility.

Building Centrosomes

The centrosome is a non-membrane bound organelle that serves as the main microtubule (MT) organizing center of most animal cells. Centrosomes function to initiate and maintain cell polarity, guide cell migration, direct intracellular cargos, and properly distribute other organelles. In mitosis, centrosomes are critical for accurate construction of the mitotic spindle to ensure faithful chromosome segregation to the two daughter cells. Thus, it is not a surprise that defects in centrosome function lead to a wide range of failures at the cellular level, which in turn, leads to tissue defects and many human diseases. The lab aims to determine how centrosomes are properly constructed from their individual parts and how centrosomes function in a wide range of cell types to avoid human diseases such as polycystic kidney disease, microcephaly, cancer and many others.

• A centrosome interactome provides insight into organelle assembly and reveals a non-duplication role for Plk4
  Galletta BJ, Fagerstrom CJ, Schoborg TA, McLamarrah TA, Ryniawec JM, Buster DW, Slep KC, Rogers GC, Rusan NM 
  Nature Communications, 7:12476
• A yeast two-hybrid approach for probing protein-protein interactions at the centrosome
  Galletta BJ, Rusan NM
  Methods in Cell Biology, 129:251-77

Bridge Proteins

For many years now, our lab has focuses on understanding how the centrosome ‘bridge protein’ Pericentrin-Like-Protein serves as both a negative and positive regulator of PCM assembly. We are specifically testing the hypothesis that PLP functions as a molecular switch to repel PCM in interphase and anchor PCM in mitosis. We aim to discover the upstream regulators of PLP and its downstream effectors.

• PLP inhibits the activity of interphase centrosomes to ensure their proper segregation in stem cells
  Lerit DA, Rusan NM.
  Journal of Cell Biology. 2013 Sep 30;202(7):1013-22
• Interphase centrosome organization by the PLP-Cnn scaffold is required for centrosome function
  Lerit DA, Jordan HA, Poulton JS, Fagerstrom CJ, Galletta BJ, Peifer M, Rusan NM
  Journal of Cell Biology, 210(1):79-97
• Bridging centrioles and PCM in proper space and time
  Varadarajan R, Rusan NM
  Essays in Biochemistry, 7;62(6):793-801

Brain Development

An intimate link between centrosome function and neurogenesis is revealed by the identification of many genes with centrosome-associated functions mutated in microcephaly disorders. Consistent with the major role of the centrosome in mitosis, mutations in these centrosome-related microcephaly (CRM) genes are thought to affect neurogenesis by depleting the pool of neural progenitor cells, primarily through apoptosis as a consequence of mitotic failure, or premature differentiation as a consequence of cell cycle delay and randomization of spindle orientation. However, as suggested by the wide range of microcephaly phenotypes and the multifunctional nature of many CRM proteins, this picture of CRM gene function is incomplete. Our lab is investigating CRMs using Drosophila as a model system.

CRMs and Asymmetric Division

Our lab has been interested in neural stem cell division regulation and brain development for 10 years now. We have focused on several genes critical for mitotic spindle development and centrosome function. Most prominently, we have studies the role of Abnormal Spindles (Asp) and the CRMs Pericentrin- Like-Protein (PLP).

• Micro-computed tomography as a platform for exploring Drosophila development
  Schoborg TA, Smith SL, Smith LN, Morris HD, Rusan NM
  Development, 176685
• Same but Different – Pleiotropy in Centrosome-Related Microcephaly.
  O’Neill R, Schoborg TA, Rusan NM
  Molecular Biology of the Cell, 1;29(3):241-246
• An Asp-CaM complex is required for Centrosome-Pole Cohesion and Centrosome Inheritance in Neural Stem Cells
  Schoborg TA, Zajac AL, Fagerstrom CJ, Guillen RX, Rusan NM
  Journal of Cell Biology, 211(5):987-98
• Live imaging of Drosophila larval neuroblasts
  Lerit DA, Plevock KM, Rusan NM
  Journal of Visualized Experiments. 2014 Jul 7;(89)
• PLP inhibits the activity of interphase centrosomes to ensure their proper segregation in stem cells
  Lerit DA, Rusan NM.
  Journal of Cell Biology. 2013 Sep 30;202(7):1013-22

Male Fertility

Inheritance of genetic material from males in humans and many other animals requires flagellated motile sperm. The construction of sperm involves major, cell wide rearrangements of many complex cellular components including the nucleus, mitochondria, ER/golgi and the cytoskeleton. Failure in any of these rearrangements can result in sperm incapable of performing their function and subfertility in affected individuals.

Basal bodies in sperm and Head-Tail linkage

Our lab is interested in the proper formation of basal bodies (centriole) that ensures the nucleation and anchoring of cilia. In recent years, we have focused on sperm basal bodies and how they properly attach to haploid nuclei.

A major step in the assembly of a flagellated sperm is the establishment of a tight connection between the head, which contains the genetic material, and the tail, which provides the force for swimming. Failure to establish the connection can result in decapitated sperm and reduced fertility. At the center of this connection lies the centriole with one end anchored firmly at the nuclear envelope and the other end serving to root and template the microtubule based axoneme of the cilia/flagella that makes up the tail.

• Sperm Head-Tail Linkage Requires Restriction of Pericentriolar Material to the Proximal Centriole End
  Galletta BJ*, Ortega JM, Smith SL, Fagerstrom CJ, Fear JM, Mahadevaraju S, Oliver B, Rusan NM* (* co-corresponding authors)
  Development Cell. 2020 53(1):86-101
• Asterless is required for centriole length control and sperm development
  Galletta BJ, Jacobs KC, Fagerstrom CJ, Rusan NM 
  Journal of Cell Biology, 23;213(4):435-50
• Drosophila Pericentrin requires interaction with Calmodulin for its function at centrosomes and neuronal basal bodies, but not at sperm basal bodies
  Galletta BJ#, Guillen RX#, Fagerstrom CJ, Brownlee CW, Lerit DA, Megraw TL, Rogers GC, Rusan NM.
  (# co-first authors)
  Molecular Biology of the Cell, 2014 Sep 15;25(18):2682-94