Biology subjects

Tembo, M.

Publications and source records attributed to Tembo, M..

6 recordsLinked to original sources

bioRxiv2022

PIP2 and Ca2+ regulation of TMEM16A currents in excised inside-out patches

The Ca2+ activated Cl- channel formed by transmembrane member 16A (TMEM16A) is broadly expressed and regulates diverse processes. In addition to Ca2+, TMEM16A channels require the acidic phospholipid phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) to open. Like other channels regulated by PI(4,5)P2, TMEM16A-conducted currents recorded in excised patches slowly decay overtime. Here we assessed how intracellular Ca2+ alters the rate of this current rundown, using the channels endogenously expressed in oocytes from the African clawed frog, Xenopus laevis. We found that in excised, inside-out patches, the concentration of applied Ca2+ alters the rate of rundown, with high Ca2+ concentrations speeding rundown by activating membrane associated phospholipase C (PLC). Together, these results clarify our understanding of how Ca2+ regulates both TMEM16A directly, and targets PLC to regulate the membrane PI(4,5)P2 content.

physiology↗

bioRxiv2020

Inhibiting actin polymerization does not prevent the fast block to polyspermy in the African clawed frog, Xenopus laevis

Fertilization of an egg by more than one sperm presents one of the earliest and most prevalent obstacles to successful reproduction. As such, eggs employ multiple mechanisms to prevent sperm entry into the nascent zygote. The fast block to polyspermy is a depolarization of the egg membrane initiated by sperm entry and is employed by diverse external fertilizers including frogs and sea urchins. For some external fertilizers, sperm entry is associated with actin polymerization during the initiation of the fast block. We therefore sought to determine whether the fast block to polyspermy in the African clawed frog, Xenopus laevis, requires actin polymerization. Although actin polymerization is required for sperm entry into eggs from diverse external fertilizers, including sea urchins and zebrafish, here we demonstrate that actin polymerization is not required for the fast block to polyspermy in X. laevis.Competing Interest StatementThe authors have declared no competing interest.View Full Text

physiology↗

bioRxiv2020

Zinc protection of fertilized eggs is an ancient feature of sexual reproduction in animals

One of the earliest and most prevalent barriers to successful reproduction is polyspermy, or fertilization of an egg by multiple sperm. To prevent these supernumerary fertilizations, eggs have evolved multiple mechanisms. It has recently been proposed that zinc released by mammalian eggs at fertilization may block additional sperm from entering. Here, we demonstrate that eggs from amphibia and teleost fish also release zinc. Using Xenopus laevis as a model, we document that zinc reversibly blocks fertilization. Finally, we demonstrate that extracellular zinc similarly disrupts early embryonic development in eggs from diverse phyla, including: Cnidaria, Echinodermata, and Chordata. Our study reveals that a fundamental strategy protecting human eggs from fertilization by multiple sperm may have evolved more than 650 million years ago.Competing Interest StatementThe authors have declared no competing interest.View Full Text

physiology↗

bioRxiv2019

Cassava planting material movement and grower behaviour in Zambia: implications for disease management

Cassava is an important food crop for most small-holder growers across sub-Saharan Africa, where production is largely limited by the presence of two viral diseases: cassava mosaic disease (CMD) and cassava brown streak disease (CBSD), both propagated by a vector whitefly and via human-mediated movement of infected cassava stems. Despite its importance, there is limited knowledge of growers behaviour related to planting material movement, as well as growers perception and knowledge of cassava diseases, which have major implications for disease spread and control. This study was conducted to address this knowledge gap by surveying small-holder growers in Zambia. A total of 96 subsistence cassava growers across five provinces were surveyed between 2015 and 2017. Most growers interviewed used planting materials from their own fields (94%) or those of nearby (<10 km) family and friends, although some large transactions with markets, middlemen, and NGOs occurred over longer distances. At the same time, information related to cassava diseases and uninfected planting material only reached 48% of growers, where those with access to information were more concerned about the disease. These data provide a basis for future planning of cassava clean seed systems to control virus diseases, emphasising the critical role of grower knowledge, and consequently education, in success of these systems. In particular, we highlight the importance of extension workers in this education process, as well as farmers groups and the media.

epidemiology↗

bioRxiv2018

The TMEM16A Channel Mediates the fast polyspermy block in Xenopus Laevis

In externally fertilizing animals, such as sea urchins and frogs, prolonged depolarization of the egg immediately after fertilization inhibits the entry of additional sperm - a phenomenon known as the fast block to polyspermy. In the African clawed frog, Xenopus laevis, this depolarization is driven by a Ca2+-activated Cl- efflux. Although the prominent Ca2+-activated Cl- currents generated by immature X. laevis oocytes are conducted by xTMEM16A channels, little is known about which channels contribute to fertilization-competency in mature eggs. Moreover, the gamete undergoes a gross transformation as it matures from an immature oocyte into a fertilization-competent egg. Here we report the results of our approach to identify the Ca2+-activated Cl- channel that triggers the fast block. Querying published proteomics and RNA-seq data, we identified two Ca2+-activated Cl- channels expressed in fertilization-competent X. laevis eggs: xTMEM16A and xBEST2A. Furthermore, transcripts for these channels increase in abundance during gamete maturation. To determine if either of these mediates the fast block, we characterized exogenously expressed xTMEM16A and xBEST2A using pharmacologic inhibitors. None of the inhibitors tested blocked xBEST2A currents specifically. However, Ani9 and MONNA each reduced xTMEM16A currents by more than 70%, while only nominally inhibiting those generated by xBEST2A. Using whole-cell recordings during fertilization, we found that Ani9 and MONNA effectively diminished fertilization-evoked depolarizations. These results indicate that fertilization activates TMEM16A channels in X. laevis eggs and induces the earliest known event triggered by fertilization: the fast block to polyspermy.\n\nHIGHLIGHTSO_LIProtein for the channels xBEST2A and xTMEM16A is present in X. laevis eggs.\nC_LI\n\nO_LIThe inhibitors MONNA and Ani9 effectively block xTMEM16A compared to xBEST2A.\nC_LI\n\nO_LIXenopus laevis fertilization opens TMEM16A to trigger egg depolarization.\nC_LI\n\nO_LIThe TMEM16A-mediated depolarization is critical for the fast block to polyspermy.\nC_LI

developmental biology↗

bioRxiv2017

Hammondia hammondi has a developmental program in vitro that mirrors its stringent two host life cycle

Hammondia hammondi is the nearest relative of Toxoplasma gondii, but unlike T. gondii is obligately heteroxenous. We have compared H. hammondi and T. gondii development in vitro and identified multiple H. hammondi-specific growth states. Despite replicating slower than T. gondii, H. hammondi was resistant to pH-induced tissue cyst formation early after excystation. However, in the absence of stress H. hammondi spontaneously converted to a terminally differentiated tissue cyst stage while T. gondii did not. Cultured H. hammondi could infect new host cells for up to 8 days following excystation, and this period was exploited to generate stably transgenic H. hammondi. Coupled with RNAseq analyses, our data clearly show that H. hammondi zoites grow as stringently regulated life stages that are fundamentally distinct from T. gondii tachyzoites and bradyzoites.

microbiology↗