Correction: Kinesin-2 transports Orco into the olfactory cilium of Drosophila melanogaster at specific developmental stages.

[This corrects the article DOI: 10.1371/journal.pgen.1009752.].

Kinesin-2 transports Orco into the olfactory cilium of Drosophila melanogaster at specific developmental stages.

The cilium, the sensing centre for the cell, displays an extensive repertoire of receptors for various cell signalling processes. The dynamic nature of ciliary signalling indicates that the ciliary entry of receptors and associated proteins must be regulated and conditional. To understand this process, we studied the ciliary localisation of the odour-receptor coreceptor (Orco), a seven-pass transmembrane protein essential for insect olfaction. Little is known about when and how Orco gets into the cilia. Here, using Drosophila melanogaster, we show that the bulk of Orco selectively enters the cilia on adult olfactory sensory neurons in two discrete, one-hour intervals after eclosion. A conditional loss of heterotrimeric kinesin-2 during this period reduces the electrophysiological response to odours and affects olfactory behaviour. We further show that Orco binds to the C-terminal tail fragments of the heterotrimeric kinesin-2 motor, which is required to transfer Orco from the ciliary base to the outer segment and maintain within an approximately four-micron stretch at the distal portion of the ciliary outer-segment. The Orco transport was not affected by the loss of critical intraflagellar transport components, IFT172/Oseg2 and IFT88/NompB, respectively, during the adult stage. These results highlight a novel developmental regulation of seven-pass transmembrane receptor transport into the cilia and indicate that ciliary signalling is both developmentally and temporally regulated.

An actomyosin clamp assembled by the Amphiphysin-Rho1-Dia/DAAM-Rok pathway reinforces somatic cell membrane folded around spermatid heads.

Membrane curvature recruits Bin-Amphiphysin-Rvs (BAR)-domain proteins and induces local F-actin assembly, which further modifies the membrane curvature and dynamics. The downstream molecular pathway in vivo is still unclear. Here, we show that a tubular endomembrane scaffold supported by contractile actomyosin stabilizes the somatic cyst cell membrane folded around rigid spermatid heads during the final stages of sperm maturation in Drosophila testis. The structure resembles an actin "basket" covering the bundle of spermatid heads. Genetic analyses suggest that the actomyosin organization is nucleated exclusively by the formins - Diaphanous and Dishevelled Associated Activator of Morphogenesis (DAAM) - downstream of Rho1, which is recruited by the BAR-domain protein Amphiphysin. Actomyosin activity at the actin basket gathers the spermatid heads into a compact bundle and resists the somatic cell invasion by intruding spermatids. These observations reveal a distinct response mechanism of actin-membrane interactions, which generates a cell-adhesion-like strategy through active clamping.

Hair Styling Procedures and Hair Morphology: A Clinico-Microscopic Comparison Study.

BACKGROUND: The present study is a cross-sectional comparison to evaluate the association between hair loss and hair structural changes (gross and microscopic), and hairstyling procedures in women. METHODS: We included 94 women; and collected data on sociodemographics, clinical history, sun-exposure, and hair-product use history. Women who reported blow drying of hair, hair straightening, use of hair iron or perming in the past 6 months were classified as cases. Age matched (±2 years) women who did not report any of the above procedures in the past 6 months were controls. The following tests were done: hair pull test; hair density assessment; hair breakage index (HBI); and microscopic examination. A logistic regression model was used for estimation of the odds ratio (OR) and 95% confidence intervals (CI). RESULTS: The mean (standard deviation [SD]) age in the case and control group was 26.4 (6.3) and 27.4 (6.3) years, respectively (P = 0.43). There was no significant difference in the mean (SD) HBI (1.05 [0.08] vs 1.07 [0.05], P = 0.22) or hair density (3.28 [0.41] vs 3.16 [0.39], P = 0.19). Cases were significantly more likely to have microscopic changes compared with controls (OR: 22.0, 95% CI: 4.3, 112.6; P < 0.001). Sun exposure for more than 3 h was significantly associated with microscopic changes (OR: 6.7, 95% CI: 1.2, 39.1; P = 0.03). CONCLUSION: Women with hairstyling procedures in the past 6 months were more likely to have microscopic changes, even though there was no difference in the hair assessment parameters. Specific guidelines on use of hairstyling procedures for Indian hair should be developed.

Atypical septate junctions maintain the somatic enclosure around maturing spermatids and prevent premature sperm release in Drosophila testis.

Tight junctions prevent paracellular flow and maintain cell polarity in an epithelium. These junctions are also required for maintaining the blood-testis barrier, which is essential for sperm differentiation. Septate junctions in insects are orthologous to the tight junctions. In Drosophila testis, major septate junction components co-localize at the interface of germline and somatic cells initially, and then condense between the two somatic cells in a cyst after germline meiosis. Their localization is extensively remodeled in subsequent stages. We find that characteristic septate junctions are formed between the somatic cyst cells at the elongated spermatid stage. Consistent with previous reports, knockdown of essential junctional components - Discs-large-1 and Neurexin-IV - during the early stages disrupted sperm differentiation beyond the spermatocyte stage. Knockdown of these proteins during the final stages of spermatid maturation caused premature release of spermatids inside the testes, resulting in partial loss of male fertility. These results indicate the importance of maintaining the integrity of the somatic enclosure during spermatid coiling and release in Drosophila testis. It also highlights the functional similarity with the tight junction proteins during mammalian spermatogenesis.This article has an associated First Person interview with the first author of the paper.

Localized, Reactive F-Actin Dynamics Prevents Abnormal Somatic Cell Penetration by Mature Spermatids.

Spermatogenesis occurs inside a somatic cell enclosure. Sperm release, the most important final step and a target for contraceptives, has been extensively studied in fixed tissue preparations. Here, we provide a time-lapse description of the release process in Drosophila testis ex vivo. We show that the spermatid tails exit the somatic enclosure and enter the testicular duct first, followed by the spermatid heads. Prior to this, individual spermatid heads attempt to invade the head cyst cell, and on each occasion they are repelled by a rapid and local F-actin polymerization response from the head cyst cell. The F-actin assembly involves N-WASp, D-WIP, and Arp2/3 complex and dissipates once the spermatid head retreats back into the fold. These findings revise the existing spermiation model in Drosophila and suggest that somatic cells can actively oppose mechanical cell invasion attempts using calibrated F-actin dynamics in situ.

Anomalies in the motion dynamics of long-flagella mutants of Chlamydomonas reinhardtii.

Chlamydomonas reinhardtii has long been used as a model organism in studies of cell motility and flagellar dynamics. The motility of the well-conserved '9+2' axoneme in its flagella remains a subject of immense curiosity. Using high-speed videography and morphological analyses, we have characterized long-flagella mutants (lf1, lf2-1, lf2-5, lf3-2, and lf4) of C. reinhardtii for biophysical parameters such as swimming velocities, waveforms, beat frequencies, and swimming trajectories. These mutants are aberrant in proteins involved in the regulation of flagellar length and bring about a phenotypic increase in this length. Our results reveal that the flagellar beat frequency and swimming velocity are negatively correlated with the length of the flagella. When compared to the wild-type, any increase in the flagellar length reduces both the swimming velocities (by 26-57%) and beat frequencies (by 8-16%). We demonstrate that with no apparent aberrations/ultrastructural deformities in the mutant axonemes, it is this increased length that has a critical role to play in the motion dynamics of C. reinhardtii cells, and, provided there are no significant changes in their flagellar proteome, any increase in this length compromises the swimming velocity either by reduction of the beat frequency or by an alteration in the waveform of the flagella.

F-actin-based extensions of the head cyst cell adhere to the maturing spermatids to maintain them in a tight bundle and prevent their premature release in Drosophila testis.

BACKGROUND: In Drosophila, all the 64 clonally derived spermatocytes differentiate in syncytium inside two somatic-origin cyst cells. They elongate to form slender spermatids, which are individualized and then released into the seminal vesicle. During individualization, differentiating spermatids are organized in a tight bundle inside the cyst, which is expected to play an important role in sperm selection. However, actual significance of this process and its underlying mechanism are unclear. RESULTS: We show that dynamic F-actin-based processes extend from the head cyst cell at the start of individualization, filling the interstitial space at the rostral ends of the maturing spermatid bundle. In addition to actin, these structures contained lamin, beta-catenin, dynamin, myosin VI and several other filopodial components. Further, pharmacological and genetic analyses showed that cytoskeletal stability and dynamin function are essential for their maintenance. Disruption of these F-actin based processes was associated with spermatid bundle disassembly and premature sperm release inside the testis. CONCLUSION: Altogether, our data suggests that the head cyst cell adheres to the maturing spermatid heads through F-actin-based extensions, thus maintaining them in a tight bundle. This is likely to regulate mature sperm release into the seminal vesicle. Overall, this process bears resemblance to mammalian spermiation.

Cytoplasmic dynein-dynactin complex is required for spermatid growth but not axoneme assembly in Drosophila.

Spermatids derived from a single gonial cell remain interconnected within a cyst and elongate by synchronized growth inside the testis in Drosophila. Cylindrical spectrin-rich elongation cones form at their distal ends during the growth. The mechanism underlying this process is poorly understood. We found that developing sperm tails were abnormally coiled at the growing ends inside the cysts in the Drosophila Dynein light chain 1 (ddlc1) hemizygous mutant testis. A quantitative assay showed that average number of elongation cones was reduced, they were increasingly deformed, and average cyst lengths were shortened in ddlc1 hemizygous testes. These phenotypes were further enhanced by additional partial reduction of Dhc64C and Glued and rescued by Myc-PIN/LC8 expression in the gonial cells in ddlc1 backgrounds. Furthermore, DDLC1, DHC, and GLUED were enriched at the distal ends of growing spermatids. Finally, ultrastructure analysis of ddlc1 testes revealed abnormally formed interspermatid membrane, but the 9 + 2 microtubule organization, the radial spoke structures, and the Dynein arms of the axoneme were normal. Together, these findings suggest that axoneme assembly and spermatid growth involve independent mechanisms in Drosophila and DDLC1 interacts with the Dynein-Dynactin complex at the distal ends of spermatids to maintain the spectrin cytoskeleton assembly and cell growth.

Drosophila KAP interacts with the kinesin II motor subunit KLP64D to assemble chordotonal sensory cilia, but not sperm tails.

BACKGROUND: Kinesin II-mediated anterograde intraflagellar transport (IFT) is essential for the assembly and maintenance of flagella and cilia in various cell types. Kinesin associated protein (KAP) is identified as the non-motor accessory subunit of Kinesin II, but its role in the corresponding motor function is not understood. RESULTS: We show that mutations in the Drosophila KAP (DmKap) gene could eliminate the sensory cilia as well as the sound-evoked potentials of Johnston's organ (JO) neurons. Ultrastructure analysis of these mutants revealed that the ciliary axonemes are absent. Mutations in Klp64D, which codes for a Kinesin II motor subunit in Drosophila, show similar ciliary defects. All these defects are rescued by exclusive expression of DmKAP and KLP64D/KIF3A in the JO neurons of respective mutants. Furthermore, reduced copy number of the DmKap gene was found to enhance the defects of hypomorphic Klp64D alleles. Unexpectedly, however, both the DmKap and the Klp64D mutant adults produce vigorously motile sperm with normal axonemes. CONCLUSIONS: KAP plays an essential role in Kinesin II function, which is required for the axoneme growth and maintenance of the cilia in Drosophila type I sensory neurons. However, the flagellar assembly in Drosophila spermatids does not require Kinesin II and is independent of IFT.