Wdr47, Camsaps, and Katanin cooperate to generate ciliary central microtubules

The axonemal central pair (CP) are non-centrosomal microtubules critical for planar ciliary beat. How they form, however, is poorly understood. Here, we show that mammalian CP formation requires Wdr47, Camsaps, and microtubule-severing activity of Katanin. Katanin severs peripheral microtubules to produce central microtubule seeds in nascent cilia. Camsaps stabilize minus ends of the seeds to facilitate microtubule outgrowth, whereas Wdr47 concentrates Camsaps into the axonemal central lumen to properly position central microtubules. Wdr47 deficiency in mouse multicilia results in complete loss of CP, rotatory beat, and primary ciliary dyskinesia. Overexpression of Camsaps or their microtubule-binding regions induces central microtubules in Wdr47−/− ependymal cells but at the expense of low efficiency, abnormal numbers, and wrong location. Katanin levels and activity also impact the central microtubule number. We propose that Wdr47, Camsaps, and Katanin function together for the generation of non-centrosomal microtubule arrays in polarized subcellular compartments.

Wdr47, Camsaps, and Katanin cooperate to generate ciliary central microtubules

The axonemal central pair (CP) are non-centrosomal microtubules critical for planar ciliary beat. How they form, however, is poorly understood. Here, we show that mammalian CP formation requires cooperative activities of Katanin, Camsaps, and Wdr47. Katanin severs peripheral microtubules to produce central microtubule seeds in nascent cilia. Camsaps stabilize minus ends of the seeds to facilitate MT outgrowth, whereas Wdr47 concentrates Camsaps into the axonemal central lumen to properly position the central microtubules. Wdr47 deficiency in mouse multicilia results in complete loss of CP, rotatory beat, and primary ciliary dyskinesia. Overexpression of Camsaps induces central microtubules in Wdr47-/- ependymal cells but at the expense of low efficiency, abnormal numbers, and wrong location, whereas overexpression of a dominant inhibitor of Katanin impairs the CP formation. We propose that Wdr47, Camsaps, and Katanin constitute a general cooperative work team for the generation of non-centrosomal MT arrays in polarized subcellular compartments.

KIF17 stabilizes microtubules and contributes to epithelial morphogenesis by acting at MT plus ends with EB1 and APC

Epithelial polarization is associated with selective stabilization and reorganization of microtubule (MT) arrays. However, upstream events and downstream consequences of MT stabilization during epithelial morphogenesis are still unclear. We show that the anterograde kinesin KIF17 localizes to MT plus ends, stabilizes MTs, and affects epithelial architecture. Targeting of KIF17 to plus ends of growing MTs requires kinesin motor activity and interaction with EB1. In turn, KIF17 participates in localizing adenomatous polyposis coli (APC) to the plus ends of a subset of MTs. We found that KIF17 affects MT dynamics, polymerization rates, and MT plus end stabilization to generate posttranslationally acetylated MTs. Depletion of KIF17 from cells growing in three-dimensional matrices results in aberrant epithelial cysts that fail to generate a single central lumen and to polarize apical markers. These findings implicate KIF17 in MT stabilization events that contribute to epithelial polarization and morphogenesis.

The Cdc42 GEF Intersectin 2 controls mitotic spindle orientation to form the lumen during epithelial morphogenesis

Epithelial organs are made of tubes and cavities lined by a monolayer of polarized cells that enclose the central lumen. Lumen formation is a crucial step in the formation of epithelial organs. The Rho guanosine triphosphatase (GTPase) Cdc42, which is a master regulator of cell polarity, regulates the formation of the central lumen in epithelial morphogenesis. However, how Cdc42 is regulated during this process is still poorly understood. Guanine nucleotide exchange factors (GEFs) control the activation of small GTPases. Using the three-dimensional Madin–Darby canine kidney model, we have identified a Cdc42-specific GEF, Intersectin 2 (ITSN2), which localizes to the centrosomes and regulates Cdc42 activation during epithelial morphogenesis. Silencing of either Cdc42 or ITSN2 disrupts the correct orientation of the mitotic spindle and normal lumen formation, suggesting a direct relationship between these processes. Furthermore, we demonstrated this direct relationship using LGN, a component of the machinery for mitotic spindle positioning, whose disruption also results in lumen formation defects.

Internal fertilization in the salp Thalia democratica

Among tunicates, gamete morphology and sperm–egg interactions have been extensively investigated in ascidians, and to a lesser extent in appendicularians and thaliaceans. Sperm–egg interaction has been studied in only one salp, Pegea socia (Bosc, 1802). To determine if the pattern of internal fertilization of P. socia is generally applicable to salps, we performed an ultrastructural study on blastozooids of Thalia democratica (Forsskål, 1775). The ovary, located in the mantle near the gut, consists of a single oocyte connected to the atrial chamber wall by a “fertilization duct”, resembling a stack of single cells without a lumen. The flagellate sperm has a long corkscrew-like head with the single mitochondrion twisted around the nucleus. Fertilization is internal, and sperm actively penetrate the atrial wall and bore through the cells of the fertilization duct. During this process, the fertilization duct shortens as the cells move apart, one to one side and the next to the other, and rejoin to form a central lumen, which contains many sperm. At the same time a few sperm reach the periovular space for fertilizing the oocyte. Comparisons with P. socia indicate that this singular mode of internal fertilization with a complex corkscrew sperm actively penetrating the fertilization duct cells, probably evolved in the salp ancestor and has been modified to some extent in various genera.

Lifelong running reduces oxidative stress and degenerative changes in the testes of mice

Regular exercise can counteract the adverse effects of aging on the musculoskeletal and cardiovascular systems. In males, the normal aging process is associated with reductions in testosterone production and impaired spermatogenesis, but the underlying mechanisms and their potential modification by exercise are unknown. Here, we report that lifelong regular exercise (running) protects the testes against the adverse effects of advancing age, and that this effect of running is associated with decreased amounts of oxidative damage to proteins, lipids, and DNA in spermatogenic and Leydig cells. Six-month-old male mice were divided into a sedentary group and a group that ran an average of 1.75 km/day, until the mice reached the age of 20 months. Seminiferous tubules of runners exhibited a full complement of cells at different stages of the spermatogenic process and a clear central lumen with large numbers of spermatozoa, in contrast to sedentary mice that exhibited disorganized spermatogenic cells and lacked spermatocytes in a central lumen. Levels of protein carbonyls, nitrotyrosine, lipid peroxidation products, and oxidatively modified DNA were significantly greater in spermatogenic and Leydig cells of sedentary mice compared with runners. These findings suggest that lifelong regular exercise suppresses aging of testes by a mechanism that involves reduced oxidative damage to spermatogenic and Leydig cells.